| anno_start anno_end anno_text entity_type sentence section | |
| 44 56 binding-site site Mechanism of extracellular ion exchange and binding-site occlusion in the sodium-calcium exchanger TITLE | |
| 74 98 sodium-calcium exchanger protein_type Mechanism of extracellular ion exchange and binding-site occlusion in the sodium-calcium exchanger TITLE | |
| 0 19 Na+/Ca2+ exchangers protein_type Na+/Ca2+ exchangers utilize the Na+ electrochemical gradient across the plasma membrane to extrude intracellular Ca2+, and play a central role in Ca2+ homeostasis. ABSTRACT | |
| 32 35 Na+ chemical Na+/Ca2+ exchangers utilize the Na+ electrochemical gradient across the plasma membrane to extrude intracellular Ca2+, and play a central role in Ca2+ homeostasis. ABSTRACT | |
| 113 117 Ca2+ chemical Na+/Ca2+ exchangers utilize the Na+ electrochemical gradient across the plasma membrane to extrude intracellular Ca2+, and play a central role in Ca2+ homeostasis. ABSTRACT | |
| 146 150 Ca2+ chemical Na+/Ca2+ exchangers utilize the Na+ electrochemical gradient across the plasma membrane to extrude intracellular Ca2+, and play a central role in Ca2+ homeostasis. ABSTRACT | |
| 92 111 structural analysis experimental_method Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 119 128 exchanger protein_type Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 134 158 Methanococcus jannaschii species Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 160 166 NCX_Mj protein Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 168 176 bound to protein_state Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 177 180 Na+ chemical Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 182 186 Ca2+ chemical Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 190 194 Sr2+ chemical Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 228 231 apo protein_state Here, we elucidate their mechanisms of extracellular ion recognition and exchange through a structural analysis of the exchanger from Methanococcus jannaschii (NCX_Mj) bound to Na+, Ca2+ or Sr2+ in various occupancies and in an apo state. ABSTRACT | |
| 130 133 Na+ chemical This analysis defines the binding mode and relative affinity of these ions, establishes the structural basis for the anticipated 3Na+:1Ca2+ exchange stoichiometry, and reveals the conformational changes at the onset of the alternating-access transport mechanism. ABSTRACT | |
| 135 139 Ca2+ chemical This analysis defines the binding mode and relative affinity of these ions, establishes the structural basis for the anticipated 3Na+:1Ca2+ exchange stoichiometry, and reveals the conformational changes at the onset of the alternating-access transport mechanism. ABSTRACT | |
| 44 80 conformational free-energy landscape evidence An independent analysis of the dynamics and conformational free-energy landscape of NCX_Mj in different ion-occupancy states, based on enhanced-sampling molecular-dynamics simulations, demonstrates that the crystal structures reflect mechanistically relevant, interconverting conformations. ABSTRACT | |
| 84 90 NCX_Mj protein An independent analysis of the dynamics and conformational free-energy landscape of NCX_Mj in different ion-occupancy states, based on enhanced-sampling molecular-dynamics simulations, demonstrates that the crystal structures reflect mechanistically relevant, interconverting conformations. ABSTRACT | |
| 104 117 ion-occupancy protein_state An independent analysis of the dynamics and conformational free-energy landscape of NCX_Mj in different ion-occupancy states, based on enhanced-sampling molecular-dynamics simulations, demonstrates that the crystal structures reflect mechanistically relevant, interconverting conformations. ABSTRACT | |
| 135 183 enhanced-sampling molecular-dynamics simulations experimental_method An independent analysis of the dynamics and conformational free-energy landscape of NCX_Mj in different ion-occupancy states, based on enhanced-sampling molecular-dynamics simulations, demonstrates that the crystal structures reflect mechanistically relevant, interconverting conformations. ABSTRACT | |
| 207 225 crystal structures evidence An independent analysis of the dynamics and conformational free-energy landscape of NCX_Mj in different ion-occupancy states, based on enhanced-sampling molecular-dynamics simulations, demonstrates that the crystal structures reflect mechanistically relevant, interconverting conformations. ABSTRACT | |
| 6 18 calculations experimental_method These calculations also reveal the mechanism by which the outward-to-inward transition is controlled by the ion-occupancy state, thereby explaining the emergence of strictly-coupled Na+/Ca2+ antiport. ABSTRACT | |
| 58 65 outward protein_state These calculations also reveal the mechanism by which the outward-to-inward transition is controlled by the ion-occupancy state, thereby explaining the emergence of strictly-coupled Na+/Ca2+ antiport. ABSTRACT | |
| 69 75 inward protein_state These calculations also reveal the mechanism by which the outward-to-inward transition is controlled by the ion-occupancy state, thereby explaining the emergence of strictly-coupled Na+/Ca2+ antiport. ABSTRACT | |
| 182 185 Na+ chemical These calculations also reveal the mechanism by which the outward-to-inward transition is controlled by the ion-occupancy state, thereby explaining the emergence of strictly-coupled Na+/Ca2+ antiport. ABSTRACT | |
| 186 190 Ca2+ chemical These calculations also reveal the mechanism by which the outward-to-inward transition is controlled by the ion-occupancy state, thereby explaining the emergence of strictly-coupled Na+/Ca2+ antiport. ABSTRACT | |
| 0 19 Na+/Ca2+ exchangers protein_type Na+/Ca2+ exchangers (NCX) play physiologically essential roles in Ca2+ signaling and homeostasis. INTRO | |
| 21 24 NCX protein_type Na+/Ca2+ exchangers (NCX) play physiologically essential roles in Ca2+ signaling and homeostasis. INTRO | |
| 66 70 Ca2+ chemical Na+/Ca2+ exchangers (NCX) play physiologically essential roles in Ca2+ signaling and homeostasis. INTRO | |
| 0 3 NCX protein_type NCX catalyzes the uphill extrusion of intracellular Ca2+ across the cell membrane, by coupling this process to the downhill permeation of Na+ into the cell, with a 3 Na+ to 1 Ca2+ stoichiometry. INTRO | |
| 52 56 Ca2+ chemical NCX catalyzes the uphill extrusion of intracellular Ca2+ across the cell membrane, by coupling this process to the downhill permeation of Na+ into the cell, with a 3 Na+ to 1 Ca2+ stoichiometry. INTRO | |
| 138 141 Na+ chemical NCX catalyzes the uphill extrusion of intracellular Ca2+ across the cell membrane, by coupling this process to the downhill permeation of Na+ into the cell, with a 3 Na+ to 1 Ca2+ stoichiometry. INTRO | |
| 166 169 Na+ chemical NCX catalyzes the uphill extrusion of intracellular Ca2+ across the cell membrane, by coupling this process to the downhill permeation of Na+ into the cell, with a 3 Na+ to 1 Ca2+ stoichiometry. INTRO | |
| 175 179 Ca2+ chemical NCX catalyzes the uphill extrusion of intracellular Ca2+ across the cell membrane, by coupling this process to the downhill permeation of Na+ into the cell, with a 3 Na+ to 1 Ca2+ stoichiometry. INTRO | |
| 17 20 NCX protein_type The mechanism of NCX proteins is therefore highly likely to be consistent with the alternating-access model of secondary-active transport. INTRO | |
| 47 50 NCX protein_type The basic functional unit for ion transport in NCX consists of ten membrane-spanning segments, comprising two homologous halves. INTRO | |
| 67 93 membrane-spanning segments structure_element The basic functional unit for ion transport in NCX consists of ten membrane-spanning segments, comprising two homologous halves. INTRO | |
| 121 127 halves structure_element The basic functional unit for ion transport in NCX consists of ten membrane-spanning segments, comprising two homologous halves. INTRO | |
| 14 20 halves structure_element Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 32 48 highly conserved protein_state Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 72 80 α-repeat structure_element Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 142 152 eukaryotic taxonomy_domain Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 153 156 NCX protein_type Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 166 172 halves structure_element Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 198 229 intracellular regulatory domain structure_element Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 240 246 absent protein_state Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 250 259 microbial taxonomy_domain Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 260 263 NCX protein_type Each of these halves contains a highly conserved region, referred to as α-repeat, known to be important for ion binding and translocation; in eukaryotic NCX, the two halves are connected by a large intracellular regulatory domain, which is absent in microbial NCX (Supplementary Fig. 1). INTRO | |
| 91 94 NCX protein_type Despite a long history of physiological and functional studies, the molecular mechanism of NCX has been elusive, owing to the lack of structural information. INTRO | |
| 29 38 structure evidence Our recent atomic-resolution structure of NCX_Mj from Methanococcus jannaschii provided the first view of the basic functional unit of an NCX protein. INTRO | |
| 42 48 NCX_Mj protein Our recent atomic-resolution structure of NCX_Mj from Methanococcus jannaschii provided the first view of the basic functional unit of an NCX protein. INTRO | |
| 54 78 Methanococcus jannaschii species Our recent atomic-resolution structure of NCX_Mj from Methanococcus jannaschii provided the first view of the basic functional unit of an NCX protein. INTRO | |
| 138 141 NCX protein_type Our recent atomic-resolution structure of NCX_Mj from Methanococcus jannaschii provided the first view of the basic functional unit of an NCX protein. INTRO | |
| 5 14 structure evidence This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 25 34 exchanger protein_type This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 41 55 outward-facing protein_state This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 95 112 ion-binding sites site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 126 134 internal site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 136 140 Sint site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 143 151 external site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 153 157 Sext site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 160 172 Ca2+-binding site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 174 177 SCa site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 183 189 middle site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 191 195 Smid site This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 241 254 occluded from protein_state This structure shows the exchanger in an outward-facing conformation and reveals four putative ion-binding sites, denominated internal (Sint), external (Sext), Ca2+-binding (SCa) and middle (Smid), clustered in the center of the protein and occluded from the solvent (Fig. 1a-b). INTRO | |
| 53 63 eukaryotic taxonomy_domain With similar ion exchange properties to those of its eukaryotic counterparts, NCX_Mj provides a compelling model system to investigate the structural basis for the specificity, stoichiometry and mechanism of the ion-exchange reaction catalyzed by NCX. INTRO | |
| 78 84 NCX_Mj protein With similar ion exchange properties to those of its eukaryotic counterparts, NCX_Mj provides a compelling model system to investigate the structural basis for the specificity, stoichiometry and mechanism of the ion-exchange reaction catalyzed by NCX. INTRO | |
| 247 250 NCX protein_type With similar ion exchange properties to those of its eukaryotic counterparts, NCX_Mj provides a compelling model system to investigate the structural basis for the specificity, stoichiometry and mechanism of the ion-exchange reaction catalyzed by NCX. INTRO | |
| 43 53 structures evidence In this study, we set out to determine the structures of outward-facing wild-type NCX_Mj in complex with Na+, Ca2+ and Sr2+, at various concentrations. INTRO | |
| 57 71 outward-facing protein_state In this study, we set out to determine the structures of outward-facing wild-type NCX_Mj in complex with Na+, Ca2+ and Sr2+, at various concentrations. INTRO | |
| 72 81 wild-type protein_state In this study, we set out to determine the structures of outward-facing wild-type NCX_Mj in complex with Na+, Ca2+ and Sr2+, at various concentrations. INTRO | |
| 82 88 NCX_Mj protein In this study, we set out to determine the structures of outward-facing wild-type NCX_Mj in complex with Na+, Ca2+ and Sr2+, at various concentrations. INTRO | |
| 89 104 in complex with protein_state In this study, we set out to determine the structures of outward-facing wild-type NCX_Mj in complex with Na+, Ca2+ and Sr2+, at various concentrations. INTRO | |
| 105 108 Na+ chemical In this study, we set out to determine the structures of outward-facing wild-type NCX_Mj in complex with Na+, Ca2+ and Sr2+, at various concentrations. INTRO | |
| 110 114 Ca2+ chemical In this study, we set out to determine the structures of outward-facing wild-type NCX_Mj in complex with Na+, Ca2+ and Sr2+, at various concentrations. INTRO | |
| 119 123 Sr2+ chemical In this study, we set out to determine the structures of outward-facing wild-type NCX_Mj in complex with Na+, Ca2+ and Sr2+, at various concentrations. INTRO | |
| 6 16 structures evidence These structures reveal the mode of recognition of these ions, their relative affinities, and the mechanism of extracellular ion exchange, for a well-defined, functional conformation in a membrane-like environment. INTRO | |
| 33 63 molecular-dynamics simulations experimental_method An independent analysis based on molecular-dynamics simulations demonstrates that the structures capture mechanistically relevant states. INTRO | |
| 86 96 structures evidence An independent analysis based on molecular-dynamics simulations demonstrates that the structures capture mechanistically relevant states. INTRO | |
| 6 18 calculations experimental_method These calculations also reveal how the ion occupancy state of the outward-facing exchanger determines the feasibility of the transition to the inward-facing conformation, thereby addressing a key outstanding question in secondary-active transport, namely how the transported substrates control the alternating-access mechanism. INTRO | |
| 66 80 outward-facing protein_state These calculations also reveal how the ion occupancy state of the outward-facing exchanger determines the feasibility of the transition to the inward-facing conformation, thereby addressing a key outstanding question in secondary-active transport, namely how the transported substrates control the alternating-access mechanism. INTRO | |
| 81 90 exchanger protein_type These calculations also reveal how the ion occupancy state of the outward-facing exchanger determines the feasibility of the transition to the inward-facing conformation, thereby addressing a key outstanding question in secondary-active transport, namely how the transported substrates control the alternating-access mechanism. INTRO | |
| 143 156 inward-facing protein_state These calculations also reveal how the ion occupancy state of the outward-facing exchanger determines the feasibility of the transition to the inward-facing conformation, thereby addressing a key outstanding question in secondary-active transport, namely how the transported substrates control the alternating-access mechanism. INTRO | |
| 14 17 Na+ chemical Extracellular Na+ binding RESULTS | |
| 27 48 central binding sites site The assignment of the four central binding sites identified in the previously reported NCX_Mj structure was hampered by the presence of both Na+ and Ca2+ in the protein crystals. RESULTS | |
| 87 93 NCX_Mj protein The assignment of the four central binding sites identified in the previously reported NCX_Mj structure was hampered by the presence of both Na+ and Ca2+ in the protein crystals. RESULTS | |
| 94 103 structure evidence The assignment of the four central binding sites identified in the previously reported NCX_Mj structure was hampered by the presence of both Na+ and Ca2+ in the protein crystals. RESULTS | |
| 141 144 Na+ chemical The assignment of the four central binding sites identified in the previously reported NCX_Mj structure was hampered by the presence of both Na+ and Ca2+ in the protein crystals. RESULTS | |
| 149 153 Ca2+ chemical The assignment of the four central binding sites identified in the previously reported NCX_Mj structure was hampered by the presence of both Na+ and Ca2+ in the protein crystals. RESULTS | |
| 169 177 crystals evidence The assignment of the four central binding sites identified in the previously reported NCX_Mj structure was hampered by the presence of both Na+ and Ca2+ in the protein crystals. RESULTS | |
| 73 76 Na+ chemical To conclusively clarify this assignment, we first set out to examine the Na+ occupancy of these sites without Ca2+. RESULTS | |
| 110 114 Ca2+ chemical To conclusively clarify this assignment, we first set out to examine the Na+ occupancy of these sites without Ca2+. RESULTS | |
| 0 8 Crystals evidence Crystals were grown in 150 mM NaCl using the lipidic cubic phase (LCP) technique. RESULTS | |
| 30 34 NaCl chemical Crystals were grown in 150 mM NaCl using the lipidic cubic phase (LCP) technique. RESULTS | |
| 45 64 lipidic cubic phase experimental_method Crystals were grown in 150 mM NaCl using the lipidic cubic phase (LCP) technique. RESULTS | |
| 66 69 LCP experimental_method Crystals were grown in 150 mM NaCl using the lipidic cubic phase (LCP) technique. RESULTS | |
| 41 44 LCP experimental_method The crystallization solutions around the LCP droplets were then slowly replaced by solutions containing different concentrations of NaCl and EGTA (Methods). RESULTS | |
| 132 136 NaCl chemical The crystallization solutions around the LCP droplets were then slowly replaced by solutions containing different concentrations of NaCl and EGTA (Methods). RESULTS | |
| 141 145 EGTA chemical The crystallization solutions around the LCP droplets were then slowly replaced by solutions containing different concentrations of NaCl and EGTA (Methods). RESULTS | |
| 0 17 X-ray diffraction experimental_method X-ray diffraction of these soaked crystals revealed a Na+-dependent variation in the electron-density distribution at sites Sext, SCa and Sint, indicating a Na+ occupancy change (Fig. 1c). RESULTS | |
| 34 42 crystals evidence X-ray diffraction of these soaked crystals revealed a Na+-dependent variation in the electron-density distribution at sites Sext, SCa and Sint, indicating a Na+ occupancy change (Fig. 1c). RESULTS | |
| 54 57 Na+ chemical X-ray diffraction of these soaked crystals revealed a Na+-dependent variation in the electron-density distribution at sites Sext, SCa and Sint, indicating a Na+ occupancy change (Fig. 1c). RESULTS | |
| 85 114 electron-density distribution evidence X-ray diffraction of these soaked crystals revealed a Na+-dependent variation in the electron-density distribution at sites Sext, SCa and Sint, indicating a Na+ occupancy change (Fig. 1c). RESULTS | |
| 124 128 Sext site X-ray diffraction of these soaked crystals revealed a Na+-dependent variation in the electron-density distribution at sites Sext, SCa and Sint, indicating a Na+ occupancy change (Fig. 1c). RESULTS | |
| 130 133 SCa site X-ray diffraction of these soaked crystals revealed a Na+-dependent variation in the electron-density distribution at sites Sext, SCa and Sint, indicating a Na+ occupancy change (Fig. 1c). RESULTS | |
| 138 142 Sint site X-ray diffraction of these soaked crystals revealed a Na+-dependent variation in the electron-density distribution at sites Sext, SCa and Sint, indicating a Na+ occupancy change (Fig. 1c). RESULTS | |
| 157 160 Na+ chemical X-ray diffraction of these soaked crystals revealed a Na+-dependent variation in the electron-density distribution at sites Sext, SCa and Sint, indicating a Na+ occupancy change (Fig. 1c). RESULTS | |
| 0 20 Occupancy refinement experimental_method Occupancy refinement indicated two Na+ ions bind to Sint and SCa at low Na+ concentrations (Fig. 1c), with a slight preference for Sint (Table 1). RESULTS | |
| 35 38 Na+ chemical Occupancy refinement indicated two Na+ ions bind to Sint and SCa at low Na+ concentrations (Fig. 1c), with a slight preference for Sint (Table 1). RESULTS | |
| 52 56 Sint site Occupancy refinement indicated two Na+ ions bind to Sint and SCa at low Na+ concentrations (Fig. 1c), with a slight preference for Sint (Table 1). RESULTS | |
| 61 64 SCa site Occupancy refinement indicated two Na+ ions bind to Sint and SCa at low Na+ concentrations (Fig. 1c), with a slight preference for Sint (Table 1). RESULTS | |
| 72 75 Na+ chemical Occupancy refinement indicated two Na+ ions bind to Sint and SCa at low Na+ concentrations (Fig. 1c), with a slight preference for Sint (Table 1). RESULTS | |
| 131 135 Sint site Occupancy refinement indicated two Na+ ions bind to Sint and SCa at low Na+ concentrations (Fig. 1c), with a slight preference for Sint (Table 1). RESULTS | |
| 19 22 Na+ chemical Binding of a third Na+ to Sext occurs at higher concentrations, as no density was observed there at 10 mM Na+ or lower (Fig. 1c); Sext is however partially occupied at 20 mM Na+, and fully occupied at 150 mM (Fig. 1c). RESULTS | |
| 26 30 Sext site Binding of a third Na+ to Sext occurs at higher concentrations, as no density was observed there at 10 mM Na+ or lower (Fig. 1c); Sext is however partially occupied at 20 mM Na+, and fully occupied at 150 mM (Fig. 1c). RESULTS | |
| 70 77 density evidence Binding of a third Na+ to Sext occurs at higher concentrations, as no density was observed there at 10 mM Na+ or lower (Fig. 1c); Sext is however partially occupied at 20 mM Na+, and fully occupied at 150 mM (Fig. 1c). RESULTS | |
| 106 109 Na+ chemical Binding of a third Na+ to Sext occurs at higher concentrations, as no density was observed there at 10 mM Na+ or lower (Fig. 1c); Sext is however partially occupied at 20 mM Na+, and fully occupied at 150 mM (Fig. 1c). RESULTS | |
| 130 134 Sext site Binding of a third Na+ to Sext occurs at higher concentrations, as no density was observed there at 10 mM Na+ or lower (Fig. 1c); Sext is however partially occupied at 20 mM Na+, and fully occupied at 150 mM (Fig. 1c). RESULTS | |
| 174 177 Na+ chemical Binding of a third Na+ to Sext occurs at higher concentrations, as no density was observed there at 10 mM Na+ or lower (Fig. 1c); Sext is however partially occupied at 20 mM Na+, and fully occupied at 150 mM (Fig. 1c). RESULTS | |
| 4 7 Na+ chemical The Na+ occupation at SCa, compounded with the expected 3Na+:1Ca2+ stoichiometry, implies our previous assignment of the Smid site must be re-evaluated. RESULTS | |
| 22 25 SCa site The Na+ occupation at SCa, compounded with the expected 3Na+:1Ca2+ stoichiometry, implies our previous assignment of the Smid site must be re-evaluated. RESULTS | |
| 57 60 Na+ chemical The Na+ occupation at SCa, compounded with the expected 3Na+:1Ca2+ stoichiometry, implies our previous assignment of the Smid site must be re-evaluated. RESULTS | |
| 62 66 Ca2+ chemical The Na+ occupation at SCa, compounded with the expected 3Na+:1Ca2+ stoichiometry, implies our previous assignment of the Smid site must be re-evaluated. RESULTS | |
| 121 125 Smid site The Na+ occupation at SCa, compounded with the expected 3Na+:1Ca2+ stoichiometry, implies our previous assignment of the Smid site must be re-evaluated. RESULTS | |
| 41 46 water chemical Indeed, two observations indicate that a water molecule rather than a Na+ ion occupies Smid, as was predicted in a recent simulation study. RESULTS | |
| 70 73 Na+ chemical Indeed, two observations indicate that a water molecule rather than a Na+ ion occupies Smid, as was predicted in a recent simulation study. RESULTS | |
| 87 91 Smid site Indeed, two observations indicate that a water molecule rather than a Na+ ion occupies Smid, as was predicted in a recent simulation study. RESULTS | |
| 122 132 simulation experimental_method Indeed, two observations indicate that a water molecule rather than a Na+ ion occupies Smid, as was predicted in a recent simulation study. RESULTS | |
| 11 27 electron density evidence First, the electron density at Smid does not depend significantly on the Na+ concentration. RESULTS | |
| 31 35 Smid site First, the electron density at Smid does not depend significantly on the Na+ concentration. RESULTS | |
| 73 76 Na+ chemical First, the electron density at Smid does not depend significantly on the Na+ concentration. RESULTS | |
| 45 49 Smid site Second, the protein coordination geometry at Smid is clearly suboptimal for Na+ (Supplementary Fig. 1d). RESULTS | |
| 76 79 Na+ chemical Second, the protein coordination geometry at Smid is clearly suboptimal for Na+ (Supplementary Fig. 1d). RESULTS | |
| 4 9 water chemical The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 22 26 Smid site The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 33 47 hydrogen-bonds bond_interaction The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 57 73 highly conserved protein_state The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 74 79 Glu54 residue_name_number The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 84 90 Glu213 residue_name_number The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 158 168 coordinate bond_interaction The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 178 181 Na+ chemical The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 190 194 Sext site The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 196 199 SCa site The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 204 208 Sint site The water molecule at Smid forms hydrogen-bonds with the highly conserved Glu54 and Glu213 (Supplementary Fig. 1d), stabilizing their orientation to properly coordinate multiple Na+ ions at Sext, SCa and Sint. RESULTS | |
| 45 50 Glu54 residue_name_number It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 55 61 Glu213 residue_name_number It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 81 87 Asp240 residue_name_number It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 102 106 Smid site It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 127 130 Asn residue_name It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 134 144 eukaryotic taxonomy_domain It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 145 148 NCX protein_type It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 159 169 protonated protein_state It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 206 216 simulation experimental_method It can be inferred from this assignment that Glu54 and Glu213 are ionized, while Asp240, which flanks Smid (and is replaced by Asn in eukaryotic NCX) would be protonated, as indicated by the abovementioned simulation study. RESULTS | |
| 0 3 Na+ chemical Na+-dependent conformational change RESULTS | |
| 4 10 NCX_Mj protein The NCX_Mj structures in various Na+ concentrations also reveal that Na+ binding to Sext is coupled to a subtle but important conformational change (Fig. 2). RESULTS | |
| 11 21 structures evidence The NCX_Mj structures in various Na+ concentrations also reveal that Na+ binding to Sext is coupled to a subtle but important conformational change (Fig. 2). RESULTS | |
| 33 36 Na+ chemical The NCX_Mj structures in various Na+ concentrations also reveal that Na+ binding to Sext is coupled to a subtle but important conformational change (Fig. 2). RESULTS | |
| 69 72 Na+ chemical The NCX_Mj structures in various Na+ concentrations also reveal that Na+ binding to Sext is coupled to a subtle but important conformational change (Fig. 2). RESULTS | |
| 84 88 Sext site The NCX_Mj structures in various Na+ concentrations also reveal that Na+ binding to Sext is coupled to a subtle but important conformational change (Fig. 2). RESULTS | |
| 5 8 Na+ chemical When Na+ binds to Sext at high concentrations, the N-terminal half of TM7 is bent into two short helices, TM7a and TM7b (Fig. 2a). RESULTS | |
| 18 22 Sext site When Na+ binds to Sext at high concentrations, the N-terminal half of TM7 is bent into two short helices, TM7a and TM7b (Fig. 2a). RESULTS | |
| 26 30 high protein_state When Na+ binds to Sext at high concentrations, the N-terminal half of TM7 is bent into two short helices, TM7a and TM7b (Fig. 2a). RESULTS | |
| 51 66 N-terminal half structure_element When Na+ binds to Sext at high concentrations, the N-terminal half of TM7 is bent into two short helices, TM7a and TM7b (Fig. 2a). RESULTS | |
| 70 73 TM7 structure_element When Na+ binds to Sext at high concentrations, the N-terminal half of TM7 is bent into two short helices, TM7a and TM7b (Fig. 2a). RESULTS | |
| 91 104 short helices structure_element When Na+ binds to Sext at high concentrations, the N-terminal half of TM7 is bent into two short helices, TM7a and TM7b (Fig. 2a). RESULTS | |
| 106 110 TM7a structure_element When Na+ binds to Sext at high concentrations, the N-terminal half of TM7 is bent into two short helices, TM7a and TM7b (Fig. 2a). RESULTS | |
| 115 119 TM7b structure_element When Na+ binds to Sext at high concentrations, the N-terminal half of TM7 is bent into two short helices, TM7a and TM7b (Fig. 2a). RESULTS | |
| 0 4 TM7b structure_element TM7b occludes the four central binding sites from the external solution, with the backbone carbonyl of Ala206 coordinating the Na+ ion (Fig. 2b-d). RESULTS | |
| 23 44 central binding sites site TM7b occludes the four central binding sites from the external solution, with the backbone carbonyl of Ala206 coordinating the Na+ ion (Fig. 2b-d). RESULTS | |
| 103 109 Ala206 residue_name_number TM7b occludes the four central binding sites from the external solution, with the backbone carbonyl of Ala206 coordinating the Na+ ion (Fig. 2b-d). RESULTS | |
| 110 122 coordinating bond_interaction TM7b occludes the four central binding sites from the external solution, with the backbone carbonyl of Ala206 coordinating the Na+ ion (Fig. 2b-d). RESULTS | |
| 127 130 Na+ chemical TM7b occludes the four central binding sites from the external solution, with the backbone carbonyl of Ala206 coordinating the Na+ ion (Fig. 2b-d). RESULTS | |
| 14 18 Sext site However, when Sext becomes empty at low Na+ concentrations, TM7a and TM7b become a continuous straight helix (Fig. 2a), and the carbonyl group of Ala206 retracts away (Fig. 2b-d). RESULTS | |
| 27 32 empty protein_state However, when Sext becomes empty at low Na+ concentrations, TM7a and TM7b become a continuous straight helix (Fig. 2a), and the carbonyl group of Ala206 retracts away (Fig. 2b-d). RESULTS | |
| 36 39 low protein_state However, when Sext becomes empty at low Na+ concentrations, TM7a and TM7b become a continuous straight helix (Fig. 2a), and the carbonyl group of Ala206 retracts away (Fig. 2b-d). RESULTS | |
| 40 43 Na+ chemical However, when Sext becomes empty at low Na+ concentrations, TM7a and TM7b become a continuous straight helix (Fig. 2a), and the carbonyl group of Ala206 retracts away (Fig. 2b-d). RESULTS | |
| 60 64 TM7a structure_element However, when Sext becomes empty at low Na+ concentrations, TM7a and TM7b become a continuous straight helix (Fig. 2a), and the carbonyl group of Ala206 retracts away (Fig. 2b-d). RESULTS | |
| 69 73 TM7b structure_element However, when Sext becomes empty at low Na+ concentrations, TM7a and TM7b become a continuous straight helix (Fig. 2a), and the carbonyl group of Ala206 retracts away (Fig. 2b-d). RESULTS | |
| 103 108 helix structure_element However, when Sext becomes empty at low Na+ concentrations, TM7a and TM7b become a continuous straight helix (Fig. 2a), and the carbonyl group of Ala206 retracts away (Fig. 2b-d). RESULTS | |
| 146 152 Ala206 residue_name_number However, when Sext becomes empty at low Na+ concentrations, TM7a and TM7b become a continuous straight helix (Fig. 2a), and the carbonyl group of Ala206 retracts away (Fig. 2b-d). RESULTS | |
| 0 4 TM7a structure_element TM7a also forms hydrophobic contacts with the C-terminal half of TM6. RESULTS | |
| 16 36 hydrophobic contacts bond_interaction TM7a also forms hydrophobic contacts with the C-terminal half of TM6. RESULTS | |
| 46 61 C-terminal half structure_element TM7a also forms hydrophobic contacts with the C-terminal half of TM6. RESULTS | |
| 65 68 TM6 structure_element TM7a also forms hydrophobic contacts with the C-terminal half of TM6. RESULTS | |
| 33 42 structure evidence These contacts are absent in the structure with Na+ at Sext, in which there is an open gap between the two helices (Fig. 2b). RESULTS | |
| 48 51 Na+ chemical These contacts are absent in the structure with Na+ at Sext, in which there is an open gap between the two helices (Fig. 2b). RESULTS | |
| 55 59 Sext site These contacts are absent in the structure with Na+ at Sext, in which there is an open gap between the two helices (Fig. 2b). RESULTS | |
| 107 114 helices structure_element These contacts are absent in the structure with Na+ at Sext, in which there is an open gap between the two helices (Fig. 2b). RESULTS | |
| 38 41 TM6 structure_element This difference is noteworthy because TM6 and TM1 are believed to undergo a sliding motion, relative to the rest of the protein, when the transporter switches to the inward-facing conformation. RESULTS | |
| 46 49 TM1 structure_element This difference is noteworthy because TM6 and TM1 are believed to undergo a sliding motion, relative to the rest of the protein, when the transporter switches to the inward-facing conformation. RESULTS | |
| 138 149 transporter protein_type This difference is noteworthy because TM6 and TM1 are believed to undergo a sliding motion, relative to the rest of the protein, when the transporter switches to the inward-facing conformation. RESULTS | |
| 166 179 inward-facing protein_state This difference is noteworthy because TM6 and TM1 are believed to undergo a sliding motion, relative to the rest of the protein, when the transporter switches to the inward-facing conformation. RESULTS | |
| 21 26 TM7ab structure_element The straightening of TM7ab also opens up a passageway from the external solution to Sext and Smid, while SCa and Sint remain occluded (Fig. 2d). RESULTS | |
| 84 88 Sext site The straightening of TM7ab also opens up a passageway from the external solution to Sext and Smid, while SCa and Sint remain occluded (Fig. 2d). RESULTS | |
| 93 97 Smid site The straightening of TM7ab also opens up a passageway from the external solution to Sext and Smid, while SCa and Sint remain occluded (Fig. 2d). RESULTS | |
| 105 108 SCa site The straightening of TM7ab also opens up a passageway from the external solution to Sext and Smid, while SCa and Sint remain occluded (Fig. 2d). RESULTS | |
| 113 117 Sint site The straightening of TM7ab also opens up a passageway from the external solution to Sext and Smid, while SCa and Sint remain occluded (Fig. 2d). RESULTS | |
| 125 133 occluded protein_state The straightening of TM7ab also opens up a passageway from the external solution to Sext and Smid, while SCa and Sint remain occluded (Fig. 2d). RESULTS | |
| 10 20 structures evidence Thus, the structures at high and low Na+ concentrations represent the outward-facing occluded and partially open states, respectively. RESULTS | |
| 24 28 high protein_state Thus, the structures at high and low Na+ concentrations represent the outward-facing occluded and partially open states, respectively. RESULTS | |
| 33 36 low protein_state Thus, the structures at high and low Na+ concentrations represent the outward-facing occluded and partially open states, respectively. RESULTS | |
| 37 40 Na+ chemical Thus, the structures at high and low Na+ concentrations represent the outward-facing occluded and partially open states, respectively. RESULTS | |
| 70 84 outward-facing protein_state Thus, the structures at high and low Na+ concentrations represent the outward-facing occluded and partially open states, respectively. RESULTS | |
| 85 93 occluded protein_state Thus, the structures at high and low Na+ concentrations represent the outward-facing occluded and partially open states, respectively. RESULTS | |
| 98 112 partially open protein_state Thus, the structures at high and low Na+ concentrations represent the outward-facing occluded and partially open states, respectively. RESULTS | |
| 47 50 Na+ chemical This conformational change is dependent on the Na+ occupancy of Sext and occurs when Na+ already occupies Sint and SCa. RESULTS | |
| 64 68 Sext site This conformational change is dependent on the Na+ occupancy of Sext and occurs when Na+ already occupies Sint and SCa. RESULTS | |
| 85 88 Na+ chemical This conformational change is dependent on the Na+ occupancy of Sext and occurs when Na+ already occupies Sint and SCa. RESULTS | |
| 106 110 Sint site This conformational change is dependent on the Na+ occupancy of Sext and occurs when Na+ already occupies Sint and SCa. RESULTS | |
| 115 118 SCa site This conformational change is dependent on the Na+ occupancy of Sext and occurs when Na+ already occupies Sint and SCa. RESULTS | |
| 4 41 crystallographic titration experiment experimental_method Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 61 65 K1/2 evidence Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 74 77 Na+ chemical Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 145 149 Sext site Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 153 171 partially occupied protein_state Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 180 186 NCX_Mj protein Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 187 194 crystal evidence Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 220 228 occluded protein_state Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 233 247 partially open protein_state Our crystallographic titration experiment indicates that the K1/2 of this Na+-driven conformational transition is ~20 mM. At this concentration, Sext is partially occupied and the NCX_Mj crystal is a mixture of both the occluded and partially open conformations. RESULTS | |
| 26 38 Na+ affinity evidence This structurally-derived Na+ affinity agrees well with the external Na+ concentration required for NCX activation in eukaryotes. RESULTS | |
| 69 72 Na+ chemical This structurally-derived Na+ affinity agrees well with the external Na+ concentration required for NCX activation in eukaryotes. RESULTS | |
| 100 103 NCX protein_type This structurally-derived Na+ affinity agrees well with the external Na+ concentration required for NCX activation in eukaryotes. RESULTS | |
| 118 128 eukaryotes taxonomy_domain This structurally-derived Na+ affinity agrees well with the external Na+ concentration required for NCX activation in eukaryotes. RESULTS | |
| 21 24 Na+ chemical The finding that the Na+ occupancy change from 2 to 3 ions coincides with a conformational change of the transporter also provides a rationale to the Hill coefficient of the Na+-dependent activation process in eukaryotic NCX. RESULTS | |
| 105 116 transporter protein_type The finding that the Na+ occupancy change from 2 to 3 ions coincides with a conformational change of the transporter also provides a rationale to the Hill coefficient of the Na+-dependent activation process in eukaryotic NCX. RESULTS | |
| 150 166 Hill coefficient evidence The finding that the Na+ occupancy change from 2 to 3 ions coincides with a conformational change of the transporter also provides a rationale to the Hill coefficient of the Na+-dependent activation process in eukaryotic NCX. RESULTS | |
| 174 177 Na+ chemical The finding that the Na+ occupancy change from 2 to 3 ions coincides with a conformational change of the transporter also provides a rationale to the Hill coefficient of the Na+-dependent activation process in eukaryotic NCX. RESULTS | |
| 210 220 eukaryotic taxonomy_domain The finding that the Na+ occupancy change from 2 to 3 ions coincides with a conformational change of the transporter also provides a rationale to the Hill coefficient of the Na+-dependent activation process in eukaryotic NCX. RESULTS | |
| 221 224 NCX protein_type The finding that the Na+ occupancy change from 2 to 3 ions coincides with a conformational change of the transporter also provides a rationale to the Hill coefficient of the Na+-dependent activation process in eukaryotic NCX. RESULTS | |
| 14 18 Ca2+ chemical Extracellular Ca2+ and Sr2+ binding and their competition with Na+ RESULTS | |
| 23 27 Sr2+ chemical Extracellular Ca2+ and Sr2+ binding and their competition with Na+ RESULTS | |
| 63 66 Na+ chemical Extracellular Ca2+ and Sr2+ binding and their competition with Na+ RESULTS | |
| 17 21 Ca2+ chemical To determine how Ca2+ binds to NCX_Mj and competes with Na+, we first titrated the crystals with Sr2+ (Methods). RESULTS | |
| 31 37 NCX_Mj protein To determine how Ca2+ binds to NCX_Mj and competes with Na+, we first titrated the crystals with Sr2+ (Methods). RESULTS | |
| 56 59 Na+ chemical To determine how Ca2+ binds to NCX_Mj and competes with Na+, we first titrated the crystals with Sr2+ (Methods). RESULTS | |
| 70 91 titrated the crystals experimental_method To determine how Ca2+ binds to NCX_Mj and competes with Na+, we first titrated the crystals with Sr2+ (Methods). RESULTS | |
| 97 101 Sr2+ chemical To determine how Ca2+ binds to NCX_Mj and competes with Na+, we first titrated the crystals with Sr2+ (Methods). RESULTS | |
| 0 4 Sr2+ chemical Sr2+ is transported by NCX similarly to Ca2+ , and is distinguishable from Na+ by its greater electron-density intensity. RESULTS | |
| 23 26 NCX protein_type Sr2+ is transported by NCX similarly to Ca2+ , and is distinguishable from Na+ by its greater electron-density intensity. RESULTS | |
| 40 44 Ca2+ chemical Sr2+ is transported by NCX similarly to Ca2+ , and is distinguishable from Na+ by its greater electron-density intensity. RESULTS | |
| 75 78 Na+ chemical Sr2+ is transported by NCX similarly to Ca2+ , and is distinguishable from Na+ by its greater electron-density intensity. RESULTS | |
| 94 120 electron-density intensity evidence Sr2+ is transported by NCX similarly to Ca2+ , and is distinguishable from Na+ by its greater electron-density intensity. RESULTS | |
| 0 23 Protein crystals soaked experimental_method Protein crystals soaked with 10 mM Sr2+ and 2.5 mM Na+ revealed a strong electron-density peak at site SCa, indicating binding of a single Sr2+ ion (Fig. 3a). RESULTS | |
| 35 39 Sr2+ chemical Protein crystals soaked with 10 mM Sr2+ and 2.5 mM Na+ revealed a strong electron-density peak at site SCa, indicating binding of a single Sr2+ ion (Fig. 3a). RESULTS | |
| 51 54 Na+ chemical Protein crystals soaked with 10 mM Sr2+ and 2.5 mM Na+ revealed a strong electron-density peak at site SCa, indicating binding of a single Sr2+ ion (Fig. 3a). RESULTS | |
| 73 94 electron-density peak evidence Protein crystals soaked with 10 mM Sr2+ and 2.5 mM Na+ revealed a strong electron-density peak at site SCa, indicating binding of a single Sr2+ ion (Fig. 3a). RESULTS | |
| 103 106 SCa site Protein crystals soaked with 10 mM Sr2+ and 2.5 mM Na+ revealed a strong electron-density peak at site SCa, indicating binding of a single Sr2+ ion (Fig. 3a). RESULTS | |
| 139 143 Sr2+ chemical Protein crystals soaked with 10 mM Sr2+ and 2.5 mM Na+ revealed a strong electron-density peak at site SCa, indicating binding of a single Sr2+ ion (Fig. 3a). RESULTS | |
| 4 15 Sr2+-loaded protein_state The Sr2+-loaded NCX_Mj structure adopts the partially open conformation observed at low Na+ concentrations. RESULTS | |
| 16 22 NCX_Mj protein The Sr2+-loaded NCX_Mj structure adopts the partially open conformation observed at low Na+ concentrations. RESULTS | |
| 23 32 structure evidence The Sr2+-loaded NCX_Mj structure adopts the partially open conformation observed at low Na+ concentrations. RESULTS | |
| 44 58 partially open protein_state The Sr2+-loaded NCX_Mj structure adopts the partially open conformation observed at low Na+ concentrations. RESULTS | |
| 88 91 Na+ chemical The Sr2+-loaded NCX_Mj structure adopts the partially open conformation observed at low Na+ concentrations. RESULTS | |
| 11 15 Sr2+ chemical Binding of Sr2+, however, excludes Na+ entirely. RESULTS | |
| 35 38 Na+ chemical Binding of Sr2+, however, excludes Na+ entirely. RESULTS | |
| 0 18 Crystal titrations experimental_method Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 24 34 decreasing experimental_method Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 35 39 Sr2+ chemical Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 43 53 increasing experimental_method Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 54 57 Na+ chemical Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 76 80 Sr2+ chemical Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 94 108 outward-facing protein_state Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 109 115 NCX_Mj protein Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 170 173 Na+ chemical Crystal titrations with decreasing Sr2+ or increasing Na+ demonstrated that Sr2+ binds to the outward-facing NCX_Mj with low affinity, and that it can be out-competed by Na+ even at low concentrations (Supplementary Note 1 and Supplementary Fig. 2a-b). RESULTS | |
| 16 19 Na+ chemical Thus, in 100 mM Na+ and 10 mM Sr2+, Na+ completely replaced Sr2+ (Fig. 3a) and reverted NCX_Mj to the Na+-loaded, fully occluded state. RESULTS | |
| 30 34 Sr2+ chemical Thus, in 100 mM Na+ and 10 mM Sr2+, Na+ completely replaced Sr2+ (Fig. 3a) and reverted NCX_Mj to the Na+-loaded, fully occluded state. RESULTS | |
| 36 39 Na+ chemical Thus, in 100 mM Na+ and 10 mM Sr2+, Na+ completely replaced Sr2+ (Fig. 3a) and reverted NCX_Mj to the Na+-loaded, fully occluded state. RESULTS | |
| 60 64 Sr2+ chemical Thus, in 100 mM Na+ and 10 mM Sr2+, Na+ completely replaced Sr2+ (Fig. 3a) and reverted NCX_Mj to the Na+-loaded, fully occluded state. RESULTS | |
| 88 94 NCX_Mj protein Thus, in 100 mM Na+ and 10 mM Sr2+, Na+ completely replaced Sr2+ (Fig. 3a) and reverted NCX_Mj to the Na+-loaded, fully occluded state. RESULTS | |
| 102 112 Na+-loaded protein_state Thus, in 100 mM Na+ and 10 mM Sr2+, Na+ completely replaced Sr2+ (Fig. 3a) and reverted NCX_Mj to the Na+-loaded, fully occluded state. RESULTS | |
| 114 128 fully occluded protein_state Thus, in 100 mM Na+ and 10 mM Sr2+, Na+ completely replaced Sr2+ (Fig. 3a) and reverted NCX_Mj to the Na+-loaded, fully occluded state. RESULTS | |
| 8 29 titration experiments experimental_method Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 42 46 Ca2+ chemical Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 51 55 Sr2+ chemical Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 67 73 NCX_Mj protein Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 100 129 electron density distribution evidence Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 135 153 crystals soaked in experimental_method Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 154 158 high protein_state Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 159 163 Ca2+ chemical Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 168 171 low protein_state Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 172 175 Na+ chemical Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 192 196 Ca2+ chemical Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 209 213 Smid site Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 225 228 SCa site Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 252 255 SCa site Similar titration experiments showed that Ca2+ and Sr2+ binding to NCX_Mj are not exactly alike The electron density distribution from crystals soaked in high Ca2+ and low Na+, indicates that Ca2+ can bind to Smid as well as SCa, with a preference for SCa (Fig. 3b). RESULTS | |
| 11 15 Ca2+ chemical Binding of Ca2+ to both sites simultaneously is highly improbable due to their close proximity, and at least one water molecule can be discerned coordinating the ion (Fig. 3b). RESULTS | |
| 113 118 water chemical Binding of Ca2+ to both sites simultaneously is highly improbable due to their close proximity, and at least one water molecule can be discerned coordinating the ion (Fig. 3b). RESULTS | |
| 145 157 coordinating bond_interaction Binding of Ca2+ to both sites simultaneously is highly improbable due to their close proximity, and at least one water molecule can be discerned coordinating the ion (Fig. 3b). RESULTS | |
| 4 11 partial protein_state The partial Ca2+ occupancy at Smid is likely caused by Asp240, which flanks this site and can in principle coordinate Ca2+. RESULTS | |
| 12 16 Ca2+ chemical The partial Ca2+ occupancy at Smid is likely caused by Asp240, which flanks this site and can in principle coordinate Ca2+. RESULTS | |
| 17 26 occupancy protein_state The partial Ca2+ occupancy at Smid is likely caused by Asp240, which flanks this site and can in principle coordinate Ca2+. RESULTS | |
| 30 34 Smid site The partial Ca2+ occupancy at Smid is likely caused by Asp240, which flanks this site and can in principle coordinate Ca2+. RESULTS | |
| 55 61 Asp240 residue_name_number The partial Ca2+ occupancy at Smid is likely caused by Asp240, which flanks this site and can in principle coordinate Ca2+. RESULTS | |
| 107 117 coordinate bond_interaction The partial Ca2+ occupancy at Smid is likely caused by Asp240, which flanks this site and can in principle coordinate Ca2+. RESULTS | |
| 118 122 Ca2+ chemical The partial Ca2+ occupancy at Smid is likely caused by Asp240, which flanks this site and can in principle coordinate Ca2+. RESULTS | |
| 9 45 functional and computational studies experimental_method Previous functional and computational studies, however, indicate Asp240 becomes protonated during transport. RESULTS | |
| 65 71 Asp240 residue_name_number Previous functional and computational studies, however, indicate Asp240 becomes protonated during transport. RESULTS | |
| 80 90 protonated protein_state Previous functional and computational studies, however, indicate Asp240 becomes protonated during transport. RESULTS | |
| 16 19 NCX protein_type Indeed, in most NCX proteins Asp240 is substituted by Asn, which would likely weaken or abrogate Ca2+ binding to Smid. RESULTS | |
| 29 35 Asp240 residue_name_number Indeed, in most NCX proteins Asp240 is substituted by Asn, which would likely weaken or abrogate Ca2+ binding to Smid. RESULTS | |
| 39 50 substituted experimental_method Indeed, in most NCX proteins Asp240 is substituted by Asn, which would likely weaken or abrogate Ca2+ binding to Smid. RESULTS | |
| 54 57 Asn residue_name Indeed, in most NCX proteins Asp240 is substituted by Asn, which would likely weaken or abrogate Ca2+ binding to Smid. RESULTS | |
| 97 101 Ca2+ chemical Indeed, in most NCX proteins Asp240 is substituted by Asn, which would likely weaken or abrogate Ca2+ binding to Smid. RESULTS | |
| 113 117 Smid site Indeed, in most NCX proteins Asp240 is substituted by Asn, which would likely weaken or abrogate Ca2+ binding to Smid. RESULTS | |
| 0 3 SCa site SCa is therefore the functional Ca2+ site. RESULTS | |
| 32 41 Ca2+ site site SCa is therefore the functional Ca2+ site. RESULTS | |
| 13 17 Sr2+ chemical Similarly to Sr2+, Ca2+ binds with low affinity to outward-facing NCX_Mj and can be readily displaced by Na+ (Supplementary Note 1 and Supplementary Fig. 2c). RESULTS | |
| 19 23 Ca2+ chemical Similarly to Sr2+, Ca2+ binds with low affinity to outward-facing NCX_Mj and can be readily displaced by Na+ (Supplementary Note 1 and Supplementary Fig. 2c). RESULTS | |
| 39 47 affinity evidence Similarly to Sr2+, Ca2+ binds with low affinity to outward-facing NCX_Mj and can be readily displaced by Na+ (Supplementary Note 1 and Supplementary Fig. 2c). RESULTS | |
| 51 65 outward-facing protein_state Similarly to Sr2+, Ca2+ binds with low affinity to outward-facing NCX_Mj and can be readily displaced by Na+ (Supplementary Note 1 and Supplementary Fig. 2c). RESULTS | |
| 66 72 NCX_Mj protein Similarly to Sr2+, Ca2+ binds with low affinity to outward-facing NCX_Mj and can be readily displaced by Na+ (Supplementary Note 1 and Supplementary Fig. 2c). RESULTS | |
| 105 108 Na+ chemical Similarly to Sr2+, Ca2+ binds with low affinity to outward-facing NCX_Mj and can be readily displaced by Na+ (Supplementary Note 1 and Supplementary Fig. 2c). RESULTS | |
| 32 66 physiological and biochemical data evidence This finding is consistent with physiological and biochemical data for both eukaryotic NCX and NCX_Mj indicating that the apparent Ca2+ affinity is much lower on the extracellular than the cytoplasmic side. RESULTS | |
| 76 86 eukaryotic taxonomy_domain This finding is consistent with physiological and biochemical data for both eukaryotic NCX and NCX_Mj indicating that the apparent Ca2+ affinity is much lower on the extracellular than the cytoplasmic side. RESULTS | |
| 87 90 NCX protein_type This finding is consistent with physiological and biochemical data for both eukaryotic NCX and NCX_Mj indicating that the apparent Ca2+ affinity is much lower on the extracellular than the cytoplasmic side. RESULTS | |
| 95 101 NCX_Mj protein This finding is consistent with physiological and biochemical data for both eukaryotic NCX and NCX_Mj indicating that the apparent Ca2+ affinity is much lower on the extracellular than the cytoplasmic side. RESULTS | |
| 131 144 Ca2+ affinity evidence This finding is consistent with physiological and biochemical data for both eukaryotic NCX and NCX_Mj indicating that the apparent Ca2+ affinity is much lower on the extracellular than the cytoplasmic side. RESULTS | |
| 18 50 crystallographic titration assay experimental_method Specifically, our crystallographic titration assay indicates Ca2+ binds with sub-millimolar affinity, in good agreement with the external apparent Ca2+ affinities deduced functionally for cardiac NCX (Km ~ 0.32 mM) and NCX_Mj (Km ~ 0.175 mM). RESULTS | |
| 61 65 Ca2+ chemical Specifically, our crystallographic titration assay indicates Ca2+ binds with sub-millimolar affinity, in good agreement with the external apparent Ca2+ affinities deduced functionally for cardiac NCX (Km ~ 0.32 mM) and NCX_Mj (Km ~ 0.175 mM). RESULTS | |
| 92 100 affinity evidence Specifically, our crystallographic titration assay indicates Ca2+ binds with sub-millimolar affinity, in good agreement with the external apparent Ca2+ affinities deduced functionally for cardiac NCX (Km ~ 0.32 mM) and NCX_Mj (Km ~ 0.175 mM). RESULTS | |
| 147 162 Ca2+ affinities evidence Specifically, our crystallographic titration assay indicates Ca2+ binds with sub-millimolar affinity, in good agreement with the external apparent Ca2+ affinities deduced functionally for cardiac NCX (Km ~ 0.32 mM) and NCX_Mj (Km ~ 0.175 mM). RESULTS | |
| 196 199 NCX protein_type Specifically, our crystallographic titration assay indicates Ca2+ binds with sub-millimolar affinity, in good agreement with the external apparent Ca2+ affinities deduced functionally for cardiac NCX (Km ~ 0.32 mM) and NCX_Mj (Km ~ 0.175 mM). RESULTS | |
| 201 203 Km evidence Specifically, our crystallographic titration assay indicates Ca2+ binds with sub-millimolar affinity, in good agreement with the external apparent Ca2+ affinities deduced functionally for cardiac NCX (Km ~ 0.32 mM) and NCX_Mj (Km ~ 0.175 mM). RESULTS | |
| 219 225 NCX_Mj protein Specifically, our crystallographic titration assay indicates Ca2+ binds with sub-millimolar affinity, in good agreement with the external apparent Ca2+ affinities deduced functionally for cardiac NCX (Km ~ 0.32 mM) and NCX_Mj (Km ~ 0.175 mM). RESULTS | |
| 227 229 Km evidence Specifically, our crystallographic titration assay indicates Ca2+ binds with sub-millimolar affinity, in good agreement with the external apparent Ca2+ affinities deduced functionally for cardiac NCX (Km ~ 0.32 mM) and NCX_Mj (Km ~ 0.175 mM). RESULTS | |
| 22 51 crystal titration experiments experimental_method Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 78 91 binding sites site Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 95 109 outward-facing protein_state Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 110 116 NCX_Mj protein Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 148 152 Sint site Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 157 161 Sext site Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 166 169 Na+ chemical Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 187 190 SCa site Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 222 226 Ca2+ chemical Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 251 254 Na+ chemical Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 279 289 simulation experimental_method Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 308 312 Sr2+ chemical Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 314 318 Smid site Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 352 356 Ca2+ chemical Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 378 382 Smid site Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 410 415 water chemical Taken together, these crystal titration experiments demonstrate that the four binding sites in outward-facing NCX_Mj exhibit different specificity: Sint and Sext are Na+ specific whereas SCa, previously hypothesized to be Ca2+ specific, can also bind Na+, confirming our earlier simulation study, as well as Sr2+; Smid can also transiently accommodate Ca2+ but during transport Smid is most likely occupied by water. RESULTS | |
| 4 21 ion-binding sites site The ion-binding sites in NCX_Mj can therefore accommodate up to three Na+ ions or a single divalent ion, and occupancy by Na+ and Ca2+ (or Sr2+) are mutually exclusive, as was deduced for eukaryotic exchangers. RESULTS | |
| 25 31 NCX_Mj protein The ion-binding sites in NCX_Mj can therefore accommodate up to three Na+ ions or a single divalent ion, and occupancy by Na+ and Ca2+ (or Sr2+) are mutually exclusive, as was deduced for eukaryotic exchangers. RESULTS | |
| 70 73 Na+ chemical The ion-binding sites in NCX_Mj can therefore accommodate up to three Na+ ions or a single divalent ion, and occupancy by Na+ and Ca2+ (or Sr2+) are mutually exclusive, as was deduced for eukaryotic exchangers. RESULTS | |
| 122 125 Na+ chemical The ion-binding sites in NCX_Mj can therefore accommodate up to three Na+ ions or a single divalent ion, and occupancy by Na+ and Ca2+ (or Sr2+) are mutually exclusive, as was deduced for eukaryotic exchangers. RESULTS | |
| 130 134 Ca2+ chemical The ion-binding sites in NCX_Mj can therefore accommodate up to three Na+ ions or a single divalent ion, and occupancy by Na+ and Ca2+ (or Sr2+) are mutually exclusive, as was deduced for eukaryotic exchangers. RESULTS | |
| 139 143 Sr2+ chemical The ion-binding sites in NCX_Mj can therefore accommodate up to three Na+ ions or a single divalent ion, and occupancy by Na+ and Ca2+ (or Sr2+) are mutually exclusive, as was deduced for eukaryotic exchangers. RESULTS | |
| 188 198 eukaryotic taxonomy_domain The ion-binding sites in NCX_Mj can therefore accommodate up to three Na+ ions or a single divalent ion, and occupancy by Na+ and Ca2+ (or Sr2+) are mutually exclusive, as was deduced for eukaryotic exchangers. RESULTS | |
| 199 209 exchangers protein_type The ion-binding sites in NCX_Mj can therefore accommodate up to three Na+ ions or a single divalent ion, and occupancy by Na+ and Ca2+ (or Sr2+) are mutually exclusive, as was deduced for eukaryotic exchangers. RESULTS | |
| 2 11 structure evidence A structure of NCX_Mj without Na+ or Ca2+ bound RESULTS | |
| 15 21 NCX_Mj protein A structure of NCX_Mj without Na+ or Ca2+ bound RESULTS | |
| 22 29 without protein_state A structure of NCX_Mj without Na+ or Ca2+ bound RESULTS | |
| 30 33 Na+ chemical A structure of NCX_Mj without Na+ or Ca2+ bound RESULTS | |
| 37 41 Ca2+ chemical A structure of NCX_Mj without Na+ or Ca2+ bound RESULTS | |
| 42 47 bound protein_state A structure of NCX_Mj without Na+ or Ca2+ bound RESULTS | |
| 3 6 apo protein_state An apo state of outward-facing NCX_Mj is likely to exist transiently in physiological conditions, despite the high amounts of extracellular Na+ (~150 mM) and Ca2+ (~2 mM). RESULTS | |
| 16 30 outward-facing protein_state An apo state of outward-facing NCX_Mj is likely to exist transiently in physiological conditions, despite the high amounts of extracellular Na+ (~150 mM) and Ca2+ (~2 mM). RESULTS | |
| 31 37 NCX_Mj protein An apo state of outward-facing NCX_Mj is likely to exist transiently in physiological conditions, despite the high amounts of extracellular Na+ (~150 mM) and Ca2+ (~2 mM). RESULTS | |
| 140 143 Na+ chemical An apo state of outward-facing NCX_Mj is likely to exist transiently in physiological conditions, despite the high amounts of extracellular Na+ (~150 mM) and Ca2+ (~2 mM). RESULTS | |
| 158 162 Ca2+ chemical An apo state of outward-facing NCX_Mj is likely to exist transiently in physiological conditions, despite the high amounts of extracellular Na+ (~150 mM) and Ca2+ (~2 mM). RESULTS | |
| 29 32 apo protein_state We were able to determine an apo-state structure of NCX_Mj, by crystallizing the protein at lower pH and in the absence of Na+ (Methods). RESULTS | |
| 39 48 structure evidence We were able to determine an apo-state structure of NCX_Mj, by crystallizing the protein at lower pH and in the absence of Na+ (Methods). RESULTS | |
| 52 58 NCX_Mj protein We were able to determine an apo-state structure of NCX_Mj, by crystallizing the protein at lower pH and in the absence of Na+ (Methods). RESULTS | |
| 63 76 crystallizing experimental_method We were able to determine an apo-state structure of NCX_Mj, by crystallizing the protein at lower pH and in the absence of Na+ (Methods). RESULTS | |
| 92 100 lower pH protein_state We were able to determine an apo-state structure of NCX_Mj, by crystallizing the protein at lower pH and in the absence of Na+ (Methods). RESULTS | |
| 112 122 absence of protein_state We were able to determine an apo-state structure of NCX_Mj, by crystallizing the protein at lower pH and in the absence of Na+ (Methods). RESULTS | |
| 123 126 Na+ chemical We were able to determine an apo-state structure of NCX_Mj, by crystallizing the protein at lower pH and in the absence of Na+ (Methods). RESULTS | |
| 5 14 structure evidence This structure is similar to the partially open structure with two Na+ or either one Ca2+ or one Sr2+ ion, with two noticeable differences. RESULTS | |
| 33 47 partially open protein_state This structure is similar to the partially open structure with two Na+ or either one Ca2+ or one Sr2+ ion, with two noticeable differences. RESULTS | |
| 48 57 structure evidence This structure is similar to the partially open structure with two Na+ or either one Ca2+ or one Sr2+ ion, with two noticeable differences. RESULTS | |
| 67 70 Na+ chemical This structure is similar to the partially open structure with two Na+ or either one Ca2+ or one Sr2+ ion, with two noticeable differences. RESULTS | |
| 85 89 Ca2+ chemical This structure is similar to the partially open structure with two Na+ or either one Ca2+ or one Sr2+ ion, with two noticeable differences. RESULTS | |
| 97 101 Sr2+ chemical This structure is similar to the partially open structure with two Na+ or either one Ca2+ or one Sr2+ ion, with two noticeable differences. RESULTS | |
| 7 12 TM7ab structure_element First, TM7ab along with the extracellular half of the TM6 and TM1 swing further away from the protein core (Fig. 3c), resulting in a slightly wider passageway into the binding sites. RESULTS | |
| 28 46 extracellular half structure_element First, TM7ab along with the extracellular half of the TM6 and TM1 swing further away from the protein core (Fig. 3c), resulting in a slightly wider passageway into the binding sites. RESULTS | |
| 54 57 TM6 structure_element First, TM7ab along with the extracellular half of the TM6 and TM1 swing further away from the protein core (Fig. 3c), resulting in a slightly wider passageway into the binding sites. RESULTS | |
| 62 65 TM1 structure_element First, TM7ab along with the extracellular half of the TM6 and TM1 swing further away from the protein core (Fig. 3c), resulting in a slightly wider passageway into the binding sites. RESULTS | |
| 168 181 binding sites site First, TM7ab along with the extracellular half of the TM6 and TM1 swing further away from the protein core (Fig. 3c), resulting in a slightly wider passageway into the binding sites. RESULTS | |
| 8 13 Glu54 residue_name_number Second, Glu54 and Glu213 side chains rotate away from the binding sites and appear to form hydrogen-bonds with residues involved in ion coordination in the fully Na+-loaded structure (Fig. 3d). RESULTS | |
| 18 24 Glu213 residue_name_number Second, Glu54 and Glu213 side chains rotate away from the binding sites and appear to form hydrogen-bonds with residues involved in ion coordination in the fully Na+-loaded structure (Fig. 3d). RESULTS | |
| 58 71 binding sites site Second, Glu54 and Glu213 side chains rotate away from the binding sites and appear to form hydrogen-bonds with residues involved in ion coordination in the fully Na+-loaded structure (Fig. 3d). RESULTS | |
| 91 105 hydrogen-bonds bond_interaction Second, Glu54 and Glu213 side chains rotate away from the binding sites and appear to form hydrogen-bonds with residues involved in ion coordination in the fully Na+-loaded structure (Fig. 3d). RESULTS | |
| 132 148 ion coordination bond_interaction Second, Glu54 and Glu213 side chains rotate away from the binding sites and appear to form hydrogen-bonds with residues involved in ion coordination in the fully Na+-loaded structure (Fig. 3d). RESULTS | |
| 156 172 fully Na+-loaded protein_state Second, Glu54 and Glu213 side chains rotate away from the binding sites and appear to form hydrogen-bonds with residues involved in ion coordination in the fully Na+-loaded structure (Fig. 3d). RESULTS | |
| 173 182 structure evidence Second, Glu54 and Glu213 side chains rotate away from the binding sites and appear to form hydrogen-bonds with residues involved in ion coordination in the fully Na+-loaded structure (Fig. 3d). RESULTS | |
| 13 26 binding sites site Although the binding sites are thus fully accessible to the external solution (Fig. 3e), the lack of electron density therein indicates no ions or ordered solvent molecules. RESULTS | |
| 36 52 fully accessible protein_state Although the binding sites are thus fully accessible to the external solution (Fig. 3e), the lack of electron density therein indicates no ions or ordered solvent molecules. RESULTS | |
| 101 117 electron density evidence Although the binding sites are thus fully accessible to the external solution (Fig. 3e), the lack of electron density therein indicates no ions or ordered solvent molecules. RESULTS | |
| 5 8 apo protein_state This apo structure might therefore represent the unloaded, open state of outward-facing NCX_Mj. RESULTS | |
| 9 18 structure evidence This apo structure might therefore represent the unloaded, open state of outward-facing NCX_Mj. RESULTS | |
| 49 57 unloaded protein_state This apo structure might therefore represent the unloaded, open state of outward-facing NCX_Mj. RESULTS | |
| 59 63 open protein_state This apo structure might therefore represent the unloaded, open state of outward-facing NCX_Mj. RESULTS | |
| 73 87 outward-facing protein_state This apo structure might therefore represent the unloaded, open state of outward-facing NCX_Mj. RESULTS | |
| 88 94 NCX_Mj protein This apo structure might therefore represent the unloaded, open state of outward-facing NCX_Mj. RESULTS | |
| 20 29 structure evidence Alternatively, this structure might capture a fully protonated state of the transporter, to which Na+ and Ca2+ cannot bind. RESULTS | |
| 46 62 fully protonated protein_state Alternatively, this structure might capture a fully protonated state of the transporter, to which Na+ and Ca2+ cannot bind. RESULTS | |
| 76 87 transporter protein_type Alternatively, this structure might capture a fully protonated state of the transporter, to which Na+ and Ca2+ cannot bind. RESULTS | |
| 98 101 Na+ chemical Alternatively, this structure might capture a fully protonated state of the transporter, to which Na+ and Ca2+ cannot bind. RESULTS | |
| 106 110 Ca2+ chemical Alternatively, this structure might capture a fully protonated state of the transporter, to which Na+ and Ca2+ cannot bind. RESULTS | |
| 49 69 computer simulations experimental_method Such interpretation would be consistent with the computer simulations reported below. RESULTS | |
| 8 24 transport assays experimental_method Indeed, transport assays of NCX_Mj have shown that even in the presence of Na+ or Ca2+, low pH inactivates the transport cycle. RESULTS | |
| 28 34 NCX_Mj protein Indeed, transport assays of NCX_Mj have shown that even in the presence of Na+ or Ca2+, low pH inactivates the transport cycle. RESULTS | |
| 63 74 presence of protein_state Indeed, transport assays of NCX_Mj have shown that even in the presence of Na+ or Ca2+, low pH inactivates the transport cycle. RESULTS | |
| 75 78 Na+ chemical Indeed, transport assays of NCX_Mj have shown that even in the presence of Na+ or Ca2+, low pH inactivates the transport cycle. RESULTS | |
| 82 86 Ca2+ chemical Indeed, transport assays of NCX_Mj have shown that even in the presence of Na+ or Ca2+, low pH inactivates the transport cycle. RESULTS | |
| 88 94 low pH protein_state Indeed, transport assays of NCX_Mj have shown that even in the presence of Na+ or Ca2+, low pH inactivates the transport cycle. RESULTS | |
| 95 106 inactivates protein_state Indeed, transport assays of NCX_Mj have shown that even in the presence of Na+ or Ca2+, low pH inactivates the transport cycle. RESULTS | |
| 54 60 NCX_Mj protein Ion occupancy determines the free-energy landscape of NCX_Mj RESULTS | |
| 5 34 secondary-active transporters protein_type That secondary-active transporters are able to harness an electrochemical gradient of one substrate to power the uphill transport of another relies on a seemingly simple principle: they must not transition between outward- and inward-open conformations unless in two precise substrate occupancy states. RESULTS | |
| 214 222 outward- protein_state That secondary-active transporters are able to harness an electrochemical gradient of one substrate to power the uphill transport of another relies on a seemingly simple principle: they must not transition between outward- and inward-open conformations unless in two precise substrate occupancy states. RESULTS | |
| 227 238 inward-open protein_state That secondary-active transporters are able to harness an electrochemical gradient of one substrate to power the uphill transport of another relies on a seemingly simple principle: they must not transition between outward- and inward-open conformations unless in two precise substrate occupancy states. RESULTS | |
| 0 3 NCX protein_type NCX must be loaded either with 3 Na+ or 1 Ca2+, and therefore functions as an antiporter; symporters, by contrast, undergo the alternating-access transition only when all substrates and coupling ions are concurrently bound, or in the apo state. RESULTS | |
| 33 36 Na+ chemical NCX must be loaded either with 3 Na+ or 1 Ca2+, and therefore functions as an antiporter; symporters, by contrast, undergo the alternating-access transition only when all substrates and coupling ions are concurrently bound, or in the apo state. RESULTS | |
| 42 46 Ca2+ chemical NCX must be loaded either with 3 Na+ or 1 Ca2+, and therefore functions as an antiporter; symporters, by contrast, undergo the alternating-access transition only when all substrates and coupling ions are concurrently bound, or in the apo state. RESULTS | |
| 78 88 antiporter protein_type NCX must be loaded either with 3 Na+ or 1 Ca2+, and therefore functions as an antiporter; symporters, by contrast, undergo the alternating-access transition only when all substrates and coupling ions are concurrently bound, or in the apo state. RESULTS | |
| 90 100 symporters protein_type NCX must be loaded either with 3 Na+ or 1 Ca2+, and therefore functions as an antiporter; symporters, by contrast, undergo the alternating-access transition only when all substrates and coupling ions are concurrently bound, or in the apo state. RESULTS | |
| 217 222 bound protein_state NCX must be loaded either with 3 Na+ or 1 Ca2+, and therefore functions as an antiporter; symporters, by contrast, undergo the alternating-access transition only when all substrates and coupling ions are concurrently bound, or in the apo state. RESULTS | |
| 234 237 apo protein_state NCX must be loaded either with 3 Na+ or 1 Ca2+, and therefore functions as an antiporter; symporters, by contrast, undergo the alternating-access transition only when all substrates and coupling ions are concurrently bound, or in the apo state. RESULTS | |
| 64 100 conformational free-energy landscape evidence To examine this central question, we sought to characterize the conformational free-energy landscape of NCX_Mj and to examine its dependence on the ion-occupancy state, using molecular dynamics (MD) simulations. RESULTS | |
| 104 110 NCX_Mj protein To examine this central question, we sought to characterize the conformational free-energy landscape of NCX_Mj and to examine its dependence on the ion-occupancy state, using molecular dynamics (MD) simulations. RESULTS | |
| 175 193 molecular dynamics experimental_method To examine this central question, we sought to characterize the conformational free-energy landscape of NCX_Mj and to examine its dependence on the ion-occupancy state, using molecular dynamics (MD) simulations. RESULTS | |
| 195 197 MD experimental_method To examine this central question, we sought to characterize the conformational free-energy landscape of NCX_Mj and to examine its dependence on the ion-occupancy state, using molecular dynamics (MD) simulations. RESULTS | |
| 199 210 simulations experimental_method To examine this central question, we sought to characterize the conformational free-energy landscape of NCX_Mj and to examine its dependence on the ion-occupancy state, using molecular dynamics (MD) simulations. RESULTS | |
| 62 71 structure evidence This computational analysis was based solely on the published structure of NCX_Mj, independently of the crystallographic studies described above. RESULTS | |
| 75 81 NCX_Mj protein This computational analysis was based solely on the published structure of NCX_Mj, independently of the crystallographic studies described above. RESULTS | |
| 104 128 crystallographic studies experimental_method This computational analysis was based solely on the published structure of NCX_Mj, independently of the crystallographic studies described above. RESULTS | |
| 44 54 structures evidence As it happens, the results confirm that the structures now available are representing interconverting states of the functional cycle of NCX_Mj, while revealing how the alternating-access mechanism is controlled by the ion-occupancy state. RESULTS | |
| 136 142 NCX_Mj protein As it happens, the results confirm that the structures now available are representing interconverting states of the functional cycle of NCX_Mj, while revealing how the alternating-access mechanism is controlled by the ion-occupancy state. RESULTS | |
| 24 38 MD simulations experimental_method A series of exploratory MD simulations was initially carried out to examine what features of the NCX_Mj structure might depend on the ion-binding sites occupancy. RESULTS | |
| 97 103 NCX_Mj protein A series of exploratory MD simulations was initially carried out to examine what features of the NCX_Mj structure might depend on the ion-binding sites occupancy. RESULTS | |
| 104 113 structure evidence A series of exploratory MD simulations was initially carried out to examine what features of the NCX_Mj structure might depend on the ion-binding sites occupancy. RESULTS | |
| 134 151 ion-binding sites site A series of exploratory MD simulations was initially carried out to examine what features of the NCX_Mj structure might depend on the ion-binding sites occupancy. RESULTS | |
| 23 32 simulated experimental_method Specifically, we first simulated the outward-occluded form, in the ion configuration we previously predicted, now confirmed by the high-Na+ crystal structure described above (Fig. 1b). RESULTS | |
| 37 53 outward-occluded protein_state Specifically, we first simulated the outward-occluded form, in the ion configuration we previously predicted, now confirmed by the high-Na+ crystal structure described above (Fig. 1b). RESULTS | |
| 131 139 high-Na+ protein_state Specifically, we first simulated the outward-occluded form, in the ion configuration we previously predicted, now confirmed by the high-Na+ crystal structure described above (Fig. 1b). RESULTS | |
| 140 157 crystal structure evidence Specifically, we first simulated the outward-occluded form, in the ion configuration we previously predicted, now confirmed by the high-Na+ crystal structure described above (Fig. 1b). RESULTS | |
| 9 12 Na+ chemical That is, Na+ ions occupy Sext, SCa, and Sint, while D240 is protonated and a water molecule occupies Smid. RESULTS | |
| 25 29 Sext site That is, Na+ ions occupy Sext, SCa, and Sint, while D240 is protonated and a water molecule occupies Smid. RESULTS | |
| 31 34 SCa site That is, Na+ ions occupy Sext, SCa, and Sint, while D240 is protonated and a water molecule occupies Smid. RESULTS | |
| 40 44 Sint site That is, Na+ ions occupy Sext, SCa, and Sint, while D240 is protonated and a water molecule occupies Smid. RESULTS | |
| 52 56 D240 residue_name_number That is, Na+ ions occupy Sext, SCa, and Sint, while D240 is protonated and a water molecule occupies Smid. RESULTS | |
| 60 70 protonated protein_state That is, Na+ ions occupy Sext, SCa, and Sint, while D240 is protonated and a water molecule occupies Smid. RESULTS | |
| 77 82 water chemical That is, Na+ ions occupy Sext, SCa, and Sint, while D240 is protonated and a water molecule occupies Smid. RESULTS | |
| 101 105 Smid site That is, Na+ ions occupy Sext, SCa, and Sint, while D240 is protonated and a water molecule occupies Smid. RESULTS | |
| 4 7 Na+ chemical The Na+ ion at Sext was then relocated from the site to the bulk solution (Methods), and this system was then allowed to evolve freely in time. RESULTS | |
| 15 19 Sext site The Na+ ion at Sext was then relocated from the site to the bulk solution (Methods), and this system was then allowed to evolve freely in time. RESULTS | |
| 4 7 Na+ chemical The Na+ ions at SCa and Sint were displaced subsequently, and an analogous simulation was then carried out. RESULTS | |
| 16 19 SCa site The Na+ ions at SCa and Sint were displaced subsequently, and an analogous simulation was then carried out. RESULTS | |
| 24 28 Sint site The Na+ ions at SCa and Sint were displaced subsequently, and an analogous simulation was then carried out. RESULTS | |
| 75 85 simulation experimental_method The Na+ ions at SCa and Sint were displaced subsequently, and an analogous simulation was then carried out. RESULTS | |
| 14 25 simulations experimental_method These initial simulations revealed noticeable changes in the transporter, consistent with those observed in the new crystal structures. RESULTS | |
| 61 72 transporter protein_type These initial simulations revealed noticeable changes in the transporter, consistent with those observed in the new crystal structures. RESULTS | |
| 116 134 crystal structures evidence These initial simulations revealed noticeable changes in the transporter, consistent with those observed in the new crystal structures. RESULTS | |
| 45 48 Na+ chemical The most notable change upon displacement of Na+ from Sext was the straightening of TM7ab (Fig. 4a). RESULTS | |
| 54 58 Sext site The most notable change upon displacement of Na+ from Sext was the straightening of TM7ab (Fig. 4a). RESULTS | |
| 84 89 TM7ab structure_element The most notable change upon displacement of Na+ from Sext was the straightening of TM7ab (Fig. 4a). RESULTS | |
| 7 10 Na+ chemical When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 20 25 bound protein_state When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 27 32 TM7ab structure_element When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 80 99 α-helical fragments structure_element When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 150 164 hydrogen-bonds bond_interaction When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 173 177 F202 residue_name_number When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 182 186 A206 residue_name_number When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 192 196 T203 residue_name_number When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 201 205 F207 residue_name_number When 3 Na+ ions are bound, TM7ab primarily folds as two distinct, non-collinear α-helical fragments, owing to the loss of the backbone carbonyl-amide hydrogen-bonds between F202 and A206, and T203 and F207 (Fig. 4b). RESULTS | |
| 25 29 Sext site This distortion occludes Sext from the exterior (Fig. 4d, 4h-i) and appears to be induced by the Na+ ion itself, which pulls the carbonyl group of A206 into its coordination sphere (Fig. 4g). RESULTS | |
| 97 100 Na+ chemical This distortion occludes Sext from the exterior (Fig. 4d, 4h-i) and appears to be induced by the Na+ ion itself, which pulls the carbonyl group of A206 into its coordination sphere (Fig. 4g). RESULTS | |
| 147 151 A206 residue_name_number This distortion occludes Sext from the exterior (Fig. 4d, 4h-i) and appears to be induced by the Na+ ion itself, which pulls the carbonyl group of A206 into its coordination sphere (Fig. 4g). RESULTS | |
| 5 9 Sext site With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 10 15 empty protein_state With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 26 31 TM7ab structure_element With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 50 57 α-helix structure_element With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 111 114 TM3 structure_element With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 119 122 TM7 structure_element With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 145 150 water chemical With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 202 206 Sext site With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 233 244 transporter protein_type With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 248 266 no longer occluded protein_state With Sext empty, however, TM7ab forms a canonical α-helix (Fig. 4a-b, 4g), thereby creating an opening between TM3 and TM7, which in turn allows water molecules from the external solution to reach into Sext (Fig. 4e, 4h-i), i.e. the transporter is no longer occluded. RESULTS | |
| 16 19 Na+ chemical Displacement of Na+ from SCa and Sint induces further changes (Fig. 4c). RESULTS | |
| 25 28 SCa site Displacement of Na+ from SCa and Sint induces further changes (Fig. 4c). RESULTS | |
| 33 37 Sint site Displacement of Na+ from SCa and Sint induces further changes (Fig. 4c). RESULTS | |
| 55 58 TM7 structure_element The most noticeable is an increased separation between TM7 and TM2 (Fig. 4f), previously brought together by concurrent backbone interactions with the Na+ ion at SCa (Fig. 4d-e). RESULTS | |
| 63 66 TM2 structure_element The most noticeable is an increased separation between TM7 and TM2 (Fig. 4f), previously brought together by concurrent backbone interactions with the Na+ ion at SCa (Fig. 4d-e). RESULTS | |
| 151 154 Na+ chemical The most noticeable is an increased separation between TM7 and TM2 (Fig. 4f), previously brought together by concurrent backbone interactions with the Na+ ion at SCa (Fig. 4d-e). RESULTS | |
| 162 165 SCa site The most noticeable is an increased separation between TM7 and TM2 (Fig. 4f), previously brought together by concurrent backbone interactions with the Na+ ion at SCa (Fig. 4d-e). RESULTS | |
| 0 3 TM1 structure_element TM1 and TM6 also slide further towards the membrane center, relative to the outward-occluded state (Fig. 4c). RESULTS | |
| 8 11 TM6 structure_element TM1 and TM6 also slide further towards the membrane center, relative to the outward-occluded state (Fig. 4c). RESULTS | |
| 76 92 outward-occluded protein_state TM1 and TM6 also slide further towards the membrane center, relative to the outward-occluded state (Fig. 4c). RESULTS | |
| 38 53 aqueous channel site Together, these changes open a second aqueous channel leading directly into SCa and Sint (Fig. 4f, Fig. 4h-i). RESULTS | |
| 76 79 SCa site Together, these changes open a second aqueous channel leading directly into SCa and Sint (Fig. 4f, Fig. 4h-i). RESULTS | |
| 84 88 Sint site Together, these changes open a second aqueous channel leading directly into SCa and Sint (Fig. 4f, Fig. 4h-i). RESULTS | |
| 4 15 transporter protein_type The transporter thus becomes fully outward-open. RESULTS | |
| 29 47 fully outward-open protein_state The transporter thus becomes fully outward-open. RESULTS | |
| 111 120 exchanger protein_type To more rigorously characterize the influence of the ion-occupancy state on the conformational dynamics of the exchanger, we carried out a series of enhanced-sampling MD calculations designed to reversibly simulate the transition between the outward-occluded and fully outward-open states, and thus quantify the free-energy landscape encompassing these states (Methods). RESULTS | |
| 167 182 MD calculations experimental_method To more rigorously characterize the influence of the ion-occupancy state on the conformational dynamics of the exchanger, we carried out a series of enhanced-sampling MD calculations designed to reversibly simulate the transition between the outward-occluded and fully outward-open states, and thus quantify the free-energy landscape encompassing these states (Methods). RESULTS | |
| 242 258 outward-occluded protein_state To more rigorously characterize the influence of the ion-occupancy state on the conformational dynamics of the exchanger, we carried out a series of enhanced-sampling MD calculations designed to reversibly simulate the transition between the outward-occluded and fully outward-open states, and thus quantify the free-energy landscape encompassing these states (Methods). RESULTS | |
| 263 281 fully outward-open protein_state To more rigorously characterize the influence of the ion-occupancy state on the conformational dynamics of the exchanger, we carried out a series of enhanced-sampling MD calculations designed to reversibly simulate the transition between the outward-occluded and fully outward-open states, and thus quantify the free-energy landscape encompassing these states (Methods). RESULTS | |
| 312 333 free-energy landscape evidence To more rigorously characterize the influence of the ion-occupancy state on the conformational dynamics of the exchanger, we carried out a series of enhanced-sampling MD calculations designed to reversibly simulate the transition between the outward-occluded and fully outward-open states, and thus quantify the free-energy landscape encompassing these states (Methods). RESULTS | |
| 68 71 Na+ chemical As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 75 79 Sext site As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 81 84 SCa site As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 89 93 Sint site As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 100 103 Na+ chemical As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 112 115 SCa site As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 120 124 Sint site As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 130 137 without protein_state As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 138 141 Na+ chemical As above, we initially examined three occupancy states, namely with Na+ in Sext, SCa and Sint, with Na+ only at SCa and Sint, and without Na+. RESULTS | |
| 6 18 calculations experimental_method These calculations demonstrate that the Na+ occupancy state of the transporter has a profound effect on its conformational free-energy landscape. RESULTS | |
| 40 43 Na+ chemical These calculations demonstrate that the Na+ occupancy state of the transporter has a profound effect on its conformational free-energy landscape. RESULTS | |
| 67 78 transporter protein_type These calculations demonstrate that the Na+ occupancy state of the transporter has a profound effect on its conformational free-energy landscape. RESULTS | |
| 108 144 conformational free-energy landscape evidence These calculations demonstrate that the Na+ occupancy state of the transporter has a profound effect on its conformational free-energy landscape. RESULTS | |
| 9 18 Na+ sites site When all Na+ sites are occupied, the global free-energy minimum corresponds to a conformation in which the ions are maximally coordinated by the protein (Fig. 5a, 5c); TM7ab is bent and packs closely with TM2 and TM3, and so the binding sites are occluded from the solvent (Fig. 5b). RESULTS | |
| 44 63 free-energy minimum evidence When all Na+ sites are occupied, the global free-energy minimum corresponds to a conformation in which the ions are maximally coordinated by the protein (Fig. 5a, 5c); TM7ab is bent and packs closely with TM2 and TM3, and so the binding sites are occluded from the solvent (Fig. 5b). RESULTS | |
| 168 173 TM7ab structure_element When all Na+ sites are occupied, the global free-energy minimum corresponds to a conformation in which the ions are maximally coordinated by the protein (Fig. 5a, 5c); TM7ab is bent and packs closely with TM2 and TM3, and so the binding sites are occluded from the solvent (Fig. 5b). RESULTS | |
| 205 208 TM2 structure_element When all Na+ sites are occupied, the global free-energy minimum corresponds to a conformation in which the ions are maximally coordinated by the protein (Fig. 5a, 5c); TM7ab is bent and packs closely with TM2 and TM3, and so the binding sites are occluded from the solvent (Fig. 5b). RESULTS | |
| 213 216 TM3 structure_element When all Na+ sites are occupied, the global free-energy minimum corresponds to a conformation in which the ions are maximally coordinated by the protein (Fig. 5a, 5c); TM7ab is bent and packs closely with TM2 and TM3, and so the binding sites are occluded from the solvent (Fig. 5b). RESULTS | |
| 229 242 binding sites site When all Na+ sites are occupied, the global free-energy minimum corresponds to a conformation in which the ions are maximally coordinated by the protein (Fig. 5a, 5c); TM7ab is bent and packs closely with TM2 and TM3, and so the binding sites are occluded from the solvent (Fig. 5b). RESULTS | |
| 40 51 transporter protein_type At a small energetic cost, however, the transporter can adopt a metastable ‘half-open’ conformation in which TM7ab is completely straight and Sext is open to the exterior (Fig. 5a, 5b). RESULTS | |
| 64 74 metastable protein_state At a small energetic cost, however, the transporter can adopt a metastable ‘half-open’ conformation in which TM7ab is completely straight and Sext is open to the exterior (Fig. 5a, 5b). RESULTS | |
| 76 85 half-open protein_state At a small energetic cost, however, the transporter can adopt a metastable ‘half-open’ conformation in which TM7ab is completely straight and Sext is open to the exterior (Fig. 5a, 5b). RESULTS | |
| 109 114 TM7ab structure_element At a small energetic cost, however, the transporter can adopt a metastable ‘half-open’ conformation in which TM7ab is completely straight and Sext is open to the exterior (Fig. 5a, 5b). RESULTS | |
| 142 146 Sext site At a small energetic cost, however, the transporter can adopt a metastable ‘half-open’ conformation in which TM7ab is completely straight and Sext is open to the exterior (Fig. 5a, 5b). RESULTS | |
| 150 154 open protein_state At a small energetic cost, however, the transporter can adopt a metastable ‘half-open’ conformation in which TM7ab is completely straight and Sext is open to the exterior (Fig. 5a, 5b). RESULTS | |
| 4 7 Na+ chemical The Na+ ion at Sext remains fully coordinated, but an ordered water molecule now mediates its interaction with A206:O, relieving the strain on the F202:O–A206:N hydrogen-bond (Fig. 5c). RESULTS | |
| 15 19 Sext site The Na+ ion at Sext remains fully coordinated, but an ordered water molecule now mediates its interaction with A206:O, relieving the strain on the F202:O–A206:N hydrogen-bond (Fig. 5c). RESULTS | |
| 28 45 fully coordinated protein_state The Na+ ion at Sext remains fully coordinated, but an ordered water molecule now mediates its interaction with A206:O, relieving the strain on the F202:O–A206:N hydrogen-bond (Fig. 5c). RESULTS | |
| 62 67 water chemical The Na+ ion at Sext remains fully coordinated, but an ordered water molecule now mediates its interaction with A206:O, relieving the strain on the F202:O–A206:N hydrogen-bond (Fig. 5c). RESULTS | |
| 111 115 A206 residue_name_number The Na+ ion at Sext remains fully coordinated, but an ordered water molecule now mediates its interaction with A206:O, relieving the strain on the F202:O–A206:N hydrogen-bond (Fig. 5c). RESULTS | |
| 147 151 F202 residue_name_number The Na+ ion at Sext remains fully coordinated, but an ordered water molecule now mediates its interaction with A206:O, relieving the strain on the F202:O–A206:N hydrogen-bond (Fig. 5c). RESULTS | |
| 154 158 A206 residue_name_number The Na+ ion at Sext remains fully coordinated, but an ordered water molecule now mediates its interaction with A206:O, relieving the strain on the F202:O–A206:N hydrogen-bond (Fig. 5c). RESULTS | |
| 161 174 hydrogen-bond bond_interaction The Na+ ion at Sext remains fully coordinated, but an ordered water molecule now mediates its interaction with A206:O, relieving the strain on the F202:O–A206:N hydrogen-bond (Fig. 5c). RESULTS | |
| 5 14 semi-open protein_state This semi-open conformation is nearly identical to that found to be the most probable when Na+ occupies only SCa and Sint (2 × Na+, Fig. 5a), demonstrating that binding (or release) of Na+ to Sext occurs in this metastable conformation. RESULTS | |
| 91 94 Na+ chemical This semi-open conformation is nearly identical to that found to be the most probable when Na+ occupies only SCa and Sint (2 × Na+, Fig. 5a), demonstrating that binding (or release) of Na+ to Sext occurs in this metastable conformation. RESULTS | |
| 109 112 SCa site This semi-open conformation is nearly identical to that found to be the most probable when Na+ occupies only SCa and Sint (2 × Na+, Fig. 5a), demonstrating that binding (or release) of Na+ to Sext occurs in this metastable conformation. RESULTS | |
| 117 121 Sint site This semi-open conformation is nearly identical to that found to be the most probable when Na+ occupies only SCa and Sint (2 × Na+, Fig. 5a), demonstrating that binding (or release) of Na+ to Sext occurs in this metastable conformation. RESULTS | |
| 127 130 Na+ chemical This semi-open conformation is nearly identical to that found to be the most probable when Na+ occupies only SCa and Sint (2 × Na+, Fig. 5a), demonstrating that binding (or release) of Na+ to Sext occurs in this metastable conformation. RESULTS | |
| 185 188 Na+ chemical This semi-open conformation is nearly identical to that found to be the most probable when Na+ occupies only SCa and Sint (2 × Na+, Fig. 5a), demonstrating that binding (or release) of Na+ to Sext occurs in this metastable conformation. RESULTS | |
| 192 196 Sext site This semi-open conformation is nearly identical to that found to be the most probable when Na+ occupies only SCa and Sint (2 × Na+, Fig. 5a), demonstrating that binding (or release) of Na+ to Sext occurs in this metastable conformation. RESULTS | |
| 212 222 metastable protein_state This semi-open conformation is nearly identical to that found to be the most probable when Na+ occupies only SCa and Sint (2 × Na+, Fig. 5a), demonstrating that binding (or release) of Na+ to Sext occurs in this metastable conformation. RESULTS | |
| 92 107 aqueous channel site Interestingly, this doubly occupied state can also access conformations in which the second aqueous channel mentioned above, i.e. leading to SCa between TM7 and TM2 and over the gating helices TM1 and TM6, also becomes open (Fig. 5b-c). RESULTS | |
| 141 144 SCa site Interestingly, this doubly occupied state can also access conformations in which the second aqueous channel mentioned above, i.e. leading to SCa between TM7 and TM2 and over the gating helices TM1 and TM6, also becomes open (Fig. 5b-c). RESULTS | |
| 153 156 TM7 structure_element Interestingly, this doubly occupied state can also access conformations in which the second aqueous channel mentioned above, i.e. leading to SCa between TM7 and TM2 and over the gating helices TM1 and TM6, also becomes open (Fig. 5b-c). RESULTS | |
| 161 164 TM2 structure_element Interestingly, this doubly occupied state can also access conformations in which the second aqueous channel mentioned above, i.e. leading to SCa between TM7 and TM2 and over the gating helices TM1 and TM6, also becomes open (Fig. 5b-c). RESULTS | |
| 178 192 gating helices structure_element Interestingly, this doubly occupied state can also access conformations in which the second aqueous channel mentioned above, i.e. leading to SCa between TM7 and TM2 and over the gating helices TM1 and TM6, also becomes open (Fig. 5b-c). RESULTS | |
| 193 196 TM1 structure_element Interestingly, this doubly occupied state can also access conformations in which the second aqueous channel mentioned above, i.e. leading to SCa between TM7 and TM2 and over the gating helices TM1 and TM6, also becomes open (Fig. 5b-c). RESULTS | |
| 201 204 TM6 structure_element Interestingly, this doubly occupied state can also access conformations in which the second aqueous channel mentioned above, i.e. leading to SCa between TM7 and TM2 and over the gating helices TM1 and TM6, also becomes open (Fig. 5b-c). RESULTS | |
| 219 223 open protein_state Interestingly, this doubly occupied state can also access conformations in which the second aqueous channel mentioned above, i.e. leading to SCa between TM7 and TM2 and over the gating helices TM1 and TM6, also becomes open (Fig. 5b-c). RESULTS | |
| 23 44 free-energy landscape evidence Crucially, though, the free-energy landscape for this partially occupied state demonstrates that the occluded conformation is no longer energetically feasible (Fig. 5a). RESULTS | |
| 54 72 partially occupied protein_state Crucially, though, the free-energy landscape for this partially occupied state demonstrates that the occluded conformation is no longer energetically feasible (Fig. 5a). RESULTS | |
| 101 109 occluded protein_state Crucially, though, the free-energy landscape for this partially occupied state demonstrates that the occluded conformation is no longer energetically feasible (Fig. 5a). RESULTS | |
| 34 37 Na+ chemical Displacement of the two remaining Na+ ions from SCa and Sint further reshapes the free-energy landscape of the transporter (No ions, Fig. 5a), which now can only adopt a fully open state featuring the two aqueous channels (Fig. 5b-c). RESULTS | |
| 48 51 SCa site Displacement of the two remaining Na+ ions from SCa and Sint further reshapes the free-energy landscape of the transporter (No ions, Fig. 5a), which now can only adopt a fully open state featuring the two aqueous channels (Fig. 5b-c). RESULTS | |
| 56 60 Sint site Displacement of the two remaining Na+ ions from SCa and Sint further reshapes the free-energy landscape of the transporter (No ions, Fig. 5a), which now can only adopt a fully open state featuring the two aqueous channels (Fig. 5b-c). RESULTS | |
| 82 103 free-energy landscape evidence Displacement of the two remaining Na+ ions from SCa and Sint further reshapes the free-energy landscape of the transporter (No ions, Fig. 5a), which now can only adopt a fully open state featuring the two aqueous channels (Fig. 5b-c). RESULTS | |
| 111 122 transporter protein_type Displacement of the two remaining Na+ ions from SCa and Sint further reshapes the free-energy landscape of the transporter (No ions, Fig. 5a), which now can only adopt a fully open state featuring the two aqueous channels (Fig. 5b-c). RESULTS | |
| 170 180 fully open protein_state Displacement of the two remaining Na+ ions from SCa and Sint further reshapes the free-energy landscape of the transporter (No ions, Fig. 5a), which now can only adopt a fully open state featuring the two aqueous channels (Fig. 5b-c). RESULTS | |
| 205 221 aqueous channels site Displacement of the two remaining Na+ ions from SCa and Sint further reshapes the free-energy landscape of the transporter (No ions, Fig. 5a), which now can only adopt a fully open state featuring the two aqueous channels (Fig. 5b-c). RESULTS | |
| 22 30 occluded protein_state The transition to the occluded state in this apo state is again energetically unfeasible. RESULTS | |
| 45 48 apo protein_state The transition to the occluded state in this apo state is again energetically unfeasible. RESULTS | |
| 67 71 open protein_state From a mechanistic standpoint, it is satisfying to observe how the open and semi-open states are each compatible with two different Na+ occupancies, explaining how sequential Na+ binding to energetically accessible conformations (prior to those binding events) progressively reshape the free-energy landscape of the transporter; by contrast, the occluded conformation is forbidden unless the Na+ occupancy is complete. RESULTS | |
| 76 85 semi-open protein_state From a mechanistic standpoint, it is satisfying to observe how the open and semi-open states are each compatible with two different Na+ occupancies, explaining how sequential Na+ binding to energetically accessible conformations (prior to those binding events) progressively reshape the free-energy landscape of the transporter; by contrast, the occluded conformation is forbidden unless the Na+ occupancy is complete. RESULTS | |
| 132 135 Na+ chemical From a mechanistic standpoint, it is satisfying to observe how the open and semi-open states are each compatible with two different Na+ occupancies, explaining how sequential Na+ binding to energetically accessible conformations (prior to those binding events) progressively reshape the free-energy landscape of the transporter; by contrast, the occluded conformation is forbidden unless the Na+ occupancy is complete. RESULTS | |
| 175 178 Na+ chemical From a mechanistic standpoint, it is satisfying to observe how the open and semi-open states are each compatible with two different Na+ occupancies, explaining how sequential Na+ binding to energetically accessible conformations (prior to those binding events) progressively reshape the free-energy landscape of the transporter; by contrast, the occluded conformation is forbidden unless the Na+ occupancy is complete. RESULTS | |
| 287 308 free-energy landscape evidence From a mechanistic standpoint, it is satisfying to observe how the open and semi-open states are each compatible with two different Na+ occupancies, explaining how sequential Na+ binding to energetically accessible conformations (prior to those binding events) progressively reshape the free-energy landscape of the transporter; by contrast, the occluded conformation is forbidden unless the Na+ occupancy is complete. RESULTS | |
| 316 327 transporter protein_type From a mechanistic standpoint, it is satisfying to observe how the open and semi-open states are each compatible with two different Na+ occupancies, explaining how sequential Na+ binding to energetically accessible conformations (prior to those binding events) progressively reshape the free-energy landscape of the transporter; by contrast, the occluded conformation is forbidden unless the Na+ occupancy is complete. RESULTS | |
| 346 354 occluded protein_state From a mechanistic standpoint, it is satisfying to observe how the open and semi-open states are each compatible with two different Na+ occupancies, explaining how sequential Na+ binding to energetically accessible conformations (prior to those binding events) progressively reshape the free-energy landscape of the transporter; by contrast, the occluded conformation is forbidden unless the Na+ occupancy is complete. RESULTS | |
| 392 417 Na+ occupancy is complete protein_state From a mechanistic standpoint, it is satisfying to observe how the open and semi-open states are each compatible with two different Na+ occupancies, explaining how sequential Na+ binding to energetically accessible conformations (prior to those binding events) progressively reshape the free-energy landscape of the transporter; by contrast, the occluded conformation is forbidden unless the Na+ occupancy is complete. RESULTS | |
| 41 44 Na+ chemical This processivity is logical since three Na+ ions are involved, but also implies that in the Ca2+-bound state, which includes a single ion, the transporter ought to be able to access all three major conformations, i.e. the outward-open state, in order to release (or re-bind) Ca2+, but also the occluded conformation, and thus the semi-open intermediate, in order to transition to the inward-open state. RESULTS | |
| 93 103 Ca2+-bound protein_state This processivity is logical since three Na+ ions are involved, but also implies that in the Ca2+-bound state, which includes a single ion, the transporter ought to be able to access all three major conformations, i.e. the outward-open state, in order to release (or re-bind) Ca2+, but also the occluded conformation, and thus the semi-open intermediate, in order to transition to the inward-open state. RESULTS | |
| 144 155 transporter protein_type This processivity is logical since three Na+ ions are involved, but also implies that in the Ca2+-bound state, which includes a single ion, the transporter ought to be able to access all three major conformations, i.e. the outward-open state, in order to release (or re-bind) Ca2+, but also the occluded conformation, and thus the semi-open intermediate, in order to transition to the inward-open state. RESULTS | |
| 223 235 outward-open protein_state This processivity is logical since three Na+ ions are involved, but also implies that in the Ca2+-bound state, which includes a single ion, the transporter ought to be able to access all three major conformations, i.e. the outward-open state, in order to release (or re-bind) Ca2+, but also the occluded conformation, and thus the semi-open intermediate, in order to transition to the inward-open state. RESULTS | |
| 276 280 Ca2+ chemical This processivity is logical since three Na+ ions are involved, but also implies that in the Ca2+-bound state, which includes a single ion, the transporter ought to be able to access all three major conformations, i.e. the outward-open state, in order to release (or re-bind) Ca2+, but also the occluded conformation, and thus the semi-open intermediate, in order to transition to the inward-open state. RESULTS | |
| 295 303 occluded protein_state This processivity is logical since three Na+ ions are involved, but also implies that in the Ca2+-bound state, which includes a single ion, the transporter ought to be able to access all three major conformations, i.e. the outward-open state, in order to release (or re-bind) Ca2+, but also the occluded conformation, and thus the semi-open intermediate, in order to transition to the inward-open state. RESULTS | |
| 331 340 semi-open protein_state This processivity is logical since three Na+ ions are involved, but also implies that in the Ca2+-bound state, which includes a single ion, the transporter ought to be able to access all three major conformations, i.e. the outward-open state, in order to release (or re-bind) Ca2+, but also the occluded conformation, and thus the semi-open intermediate, in order to transition to the inward-open state. RESULTS | |
| 385 396 inward-open protein_state This processivity is logical since three Na+ ions are involved, but also implies that in the Ca2+-bound state, which includes a single ion, the transporter ought to be able to access all three major conformations, i.e. the outward-open state, in order to release (or re-bind) Ca2+, but also the occluded conformation, and thus the semi-open intermediate, in order to transition to the inward-open state. RESULTS | |
| 26 28 H+ chemical By contrast, occupancy by H+, which as mentioned are not transported, might be compatible with a semi-open state as well as with the fully open conformation, but should not be conducive to occlusion. RESULTS | |
| 97 106 semi-open protein_state By contrast, occupancy by H+, which as mentioned are not transported, might be compatible with a semi-open state as well as with the fully open conformation, but should not be conducive to occlusion. RESULTS | |
| 133 143 fully open protein_state By contrast, occupancy by H+, which as mentioned are not transported, might be compatible with a semi-open state as well as with the fully open conformation, but should not be conducive to occlusion. RESULTS | |
| 42 71 enhanced-sampling simulations experimental_method To assess this hypothesis, we carried out enhanced-sampling simulations for the Ca2+ and H+-bound states of outward-facing NCX_Mj analogous to those described above for Na+ (see Supplementary Note 2 and Supplementary Fig. 3-4 for details on how the structures of the Ca2+-bound state was predicted). RESULTS | |
| 80 84 Ca2+ protein_state To assess this hypothesis, we carried out enhanced-sampling simulations for the Ca2+ and H+-bound states of outward-facing NCX_Mj analogous to those described above for Na+ (see Supplementary Note 2 and Supplementary Fig. 3-4 for details on how the structures of the Ca2+-bound state was predicted). RESULTS | |
| 89 97 H+-bound protein_state To assess this hypothesis, we carried out enhanced-sampling simulations for the Ca2+ and H+-bound states of outward-facing NCX_Mj analogous to those described above for Na+ (see Supplementary Note 2 and Supplementary Fig. 3-4 for details on how the structures of the Ca2+-bound state was predicted). RESULTS | |
| 108 122 outward-facing protein_state To assess this hypothesis, we carried out enhanced-sampling simulations for the Ca2+ and H+-bound states of outward-facing NCX_Mj analogous to those described above for Na+ (see Supplementary Note 2 and Supplementary Fig. 3-4 for details on how the structures of the Ca2+-bound state was predicted). RESULTS | |
| 123 129 NCX_Mj protein To assess this hypothesis, we carried out enhanced-sampling simulations for the Ca2+ and H+-bound states of outward-facing NCX_Mj analogous to those described above for Na+ (see Supplementary Note 2 and Supplementary Fig. 3-4 for details on how the structures of the Ca2+-bound state was predicted). RESULTS | |
| 169 172 Na+ chemical To assess this hypothesis, we carried out enhanced-sampling simulations for the Ca2+ and H+-bound states of outward-facing NCX_Mj analogous to those described above for Na+ (see Supplementary Note 2 and Supplementary Fig. 3-4 for details on how the structures of the Ca2+-bound state was predicted). RESULTS | |
| 249 259 structures evidence To assess this hypothesis, we carried out enhanced-sampling simulations for the Ca2+ and H+-bound states of outward-facing NCX_Mj analogous to those described above for Na+ (see Supplementary Note 2 and Supplementary Fig. 3-4 for details on how the structures of the Ca2+-bound state was predicted). RESULTS | |
| 267 277 Ca2+-bound protein_state To assess this hypothesis, we carried out enhanced-sampling simulations for the Ca2+ and H+-bound states of outward-facing NCX_Mj analogous to those described above for Na+ (see Supplementary Note 2 and Supplementary Fig. 3-4 for details on how the structures of the Ca2+-bound state was predicted). RESULTS | |
| 4 14 calculated experimental_method The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 15 36 free-energy landscape evidence The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 41 51 Ca2+-bound protein_state The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 52 58 NCX_Mj protein The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 103 107 Ca2+ chemical The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 149 155 NCX_Mj protein The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 176 180 Ca2+ chemical The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 186 194 occluded protein_state The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 196 206 dehydrated protein_state The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 274 283 exchanger protein_type The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 303 312 semi-open protein_state The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 317 321 open protein_state The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 356 360 Ca2+ chemical The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 373 376 Na+ chemical The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 402 425 aqueous access channels site The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 439 443 Sext site The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 448 451 SCa site The calculated free-energy landscape for Ca2+-bound NCX_Mj confirms the hypothesis outlined above (1 × Ca2+, Fig. 6a): consistent with the fact that NCX_Mj transports a single Ca2+, the occluded, dehydrated conformation is one of the major energetic minima, but clearly the exchanger can also adopt the semi-open and open states that would be required for Ca2+ release and Na+ entry, via either of the aqueous access channels that lead to Sext and SCa (Fig. 6b-c). RESULTS | |
| 13 24 protonation protein_state By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 28 33 Glu54 residue_name_number By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 38 44 Glu213 residue_name_number By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 55 63 occluded protein_state By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 133 139 NCX_Mj protein By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 159 166 protons chemical By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 176 184 H+-bound protein_state By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 204 213 exchanger protein_type By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 228 237 semi-open protein_state By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 267 273 low pH protein_state By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 275 278 apo protein_state By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 279 296 crystal structure evidence By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 302 304 H+ chemical By contrast, protonation of Glu54 and Glu213 makes the occluded conformation energetically unfeasible, consistent with the fact that NCX_Mj does not transport protons; in this H+-bound state, though, the exchanger can adopt the semi-open conformation captured in the low pH, apo crystal structure (2 × H+, Fig. 6a-c). RESULTS | |
| 21 54 systematic computational analysis experimental_method Taken together, this systematic computational analysis of outward-facing NCX_Mj clearly demonstrates that the alternating-access and ion-recognition mechanisms in this Na+/Ca2+ exchanger are coupled through the influence that the bound ions have on the free-energy landscape of the protein, which in turn determines whether or not the occluded conformation is energetically feasible. RESULTS | |
| 58 72 outward-facing protein_state Taken together, this systematic computational analysis of outward-facing NCX_Mj clearly demonstrates that the alternating-access and ion-recognition mechanisms in this Na+/Ca2+ exchanger are coupled through the influence that the bound ions have on the free-energy landscape of the protein, which in turn determines whether or not the occluded conformation is energetically feasible. RESULTS | |
| 73 79 NCX_Mj protein Taken together, this systematic computational analysis of outward-facing NCX_Mj clearly demonstrates that the alternating-access and ion-recognition mechanisms in this Na+/Ca2+ exchanger are coupled through the influence that the bound ions have on the free-energy landscape of the protein, which in turn determines whether or not the occluded conformation is energetically feasible. RESULTS | |
| 168 186 Na+/Ca2+ exchanger protein_type Taken together, this systematic computational analysis of outward-facing NCX_Mj clearly demonstrates that the alternating-access and ion-recognition mechanisms in this Na+/Ca2+ exchanger are coupled through the influence that the bound ions have on the free-energy landscape of the protein, which in turn determines whether or not the occluded conformation is energetically feasible. RESULTS | |
| 253 274 free-energy landscape evidence Taken together, this systematic computational analysis of outward-facing NCX_Mj clearly demonstrates that the alternating-access and ion-recognition mechanisms in this Na+/Ca2+ exchanger are coupled through the influence that the bound ions have on the free-energy landscape of the protein, which in turn determines whether or not the occluded conformation is energetically feasible. RESULTS | |
| 335 343 occluded protein_state Taken together, this systematic computational analysis of outward-facing NCX_Mj clearly demonstrates that the alternating-access and ion-recognition mechanisms in this Na+/Ca2+ exchanger are coupled through the influence that the bound ions have on the free-energy landscape of the protein, which in turn determines whether or not the occluded conformation is energetically feasible. RESULTS | |
| 5 13 occluded protein_state This occluded conformation, which is a necessary intermediate between the outward and inward-open states, and which entails the internal dehydration of the protein, is only attainable upon complete occupancy of the binding sites. RESULTS | |
| 74 81 outward protein_state This occluded conformation, which is a necessary intermediate between the outward and inward-open states, and which entails the internal dehydration of the protein, is only attainable upon complete occupancy of the binding sites. RESULTS | |
| 86 97 inward-open protein_state This occluded conformation, which is a necessary intermediate between the outward and inward-open states, and which entails the internal dehydration of the protein, is only attainable upon complete occupancy of the binding sites. RESULTS | |
| 137 148 dehydration protein_state This occluded conformation, which is a necessary intermediate between the outward and inward-open states, and which entails the internal dehydration of the protein, is only attainable upon complete occupancy of the binding sites. RESULTS | |
| 189 207 complete occupancy protein_state This occluded conformation, which is a necessary intermediate between the outward and inward-open states, and which entails the internal dehydration of the protein, is only attainable upon complete occupancy of the binding sites. RESULTS | |
| 215 228 binding sites site This occluded conformation, which is a necessary intermediate between the outward and inward-open states, and which entails the internal dehydration of the protein, is only attainable upon complete occupancy of the binding sites. RESULTS | |
| 78 85 outward protein_state The alternating-access hypothesis implicitly dictates that the switch between outward- and inward-open conformations of a given secondary-active transporter must not occur unless the appropriate type and number of substrates are recognized. DISCUSS | |
| 91 102 inward-open protein_state The alternating-access hypothesis implicitly dictates that the switch between outward- and inward-open conformations of a given secondary-active transporter must not occur unless the appropriate type and number of substrates are recognized. DISCUSS | |
| 128 144 secondary-active protein_state The alternating-access hypothesis implicitly dictates that the switch between outward- and inward-open conformations of a given secondary-active transporter must not occur unless the appropriate type and number of substrates are recognized. DISCUSS | |
| 145 156 transporter protein_type The alternating-access hypothesis implicitly dictates that the switch between outward- and inward-open conformations of a given secondary-active transporter must not occur unless the appropriate type and number of substrates are recognized. DISCUSS | |
| 32 43 antiporters protein_type It is however also non-trivial: antiporters, for example, do not undergo the alternating-access transition without a cargo, but this is precisely how membrane symporters reset their transport cycles. DISCUSS | |
| 150 169 membrane symporters protein_type It is however also non-trivial: antiporters, for example, do not undergo the alternating-access transition without a cargo, but this is precisely how membrane symporters reset their transport cycles. DISCUSS | |
| 35 45 antiporter protein_type Similarly puzzling is that a given antiporter will undergo this transition upon recognition of substrates of different charge, size and number. DISCUSS | |
| 67 77 antiporter protein_type Yet, when multiple species are to be co-translocated, by either an antiporter or a symporter, partial occupancies must not be conducive to the alternating-access switch. DISCUSS | |
| 83 92 symporter protein_type Yet, when multiple species are to be co-translocated, by either an antiporter or a symporter, partial occupancies must not be conducive to the alternating-access switch. DISCUSS | |
| 143 168 alternating-access switch site Yet, when multiple species are to be co-translocated, by either an antiporter or a symporter, partial occupancies must not be conducive to the alternating-access switch. DISCUSS | |
| 103 121 structural studies experimental_method Here, we have provided novel insights into this intriguing mechanism of conformational control through structural studies and quantitative molecular simulations of a Na+/Ca2+ exchanger. DISCUSS | |
| 126 160 quantitative molecular simulations experimental_method Here, we have provided novel insights into this intriguing mechanism of conformational control through structural studies and quantitative molecular simulations of a Na+/Ca2+ exchanger. DISCUSS | |
| 166 184 Na+/Ca2+ exchanger protein_type Here, we have provided novel insights into this intriguing mechanism of conformational control through structural studies and quantitative molecular simulations of a Na+/Ca2+ exchanger. DISCUSS | |
| 29 35 NCX_Mj protein Specifically, our studies of NCX_Mj reveal the mechanism of forward ion exchange (Fig. 7). DISCUSS | |
| 25 39 outward-facing protein_state The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 40 46 NCX_Mj protein The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 55 68 inward-facing protein_state The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 69 87 crystal structures evidence The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 99 117 Ca2+/H+ exchangers protein_type The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 168 171 NCX protein_type The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 209 212 TM1 structure_element The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 217 220 TM6 structure_element The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 249 260 transporter protein_type The internal symmetry of outward-facing NCX_Mj and the inward-facing crystal structures of several Ca2+/H+ exchangers indicate that the alternating-access mechanism of NCX proteins entails a sliding motion of TM1 and TM6 relative to the rest of the transporter. DISCUSS | |
| 56 76 extracellular region structure_element Here, we demonstrate that conformational changes in the extracellular region of the TM2-TM3 and TM7-TM8 bundle precede and are necessary for the transition, and are associated with ion recognition and/or release. DISCUSS | |
| 84 91 TM2-TM3 structure_element Here, we demonstrate that conformational changes in the extracellular region of the TM2-TM3 and TM7-TM8 bundle precede and are necessary for the transition, and are associated with ion recognition and/or release. DISCUSS | |
| 96 110 TM7-TM8 bundle structure_element Here, we demonstrate that conformational changes in the extracellular region of the TM2-TM3 and TM7-TM8 bundle precede and are necessary for the transition, and are associated with ion recognition and/or release. DISCUSS | |
| 48 63 N-terminal half structure_element The most apparent of these changes involves the N-terminal half of TM7 (TM7ab); together with more subtle displacements in TM2 and TM3, this change in TM7ab correlates with the opening and closing of two distinct aqueous channels leading into the ion-binding sites from the extracellular solution. DISCUSS | |
| 67 70 TM7 structure_element The most apparent of these changes involves the N-terminal half of TM7 (TM7ab); together with more subtle displacements in TM2 and TM3, this change in TM7ab correlates with the opening and closing of two distinct aqueous channels leading into the ion-binding sites from the extracellular solution. DISCUSS | |
| 72 77 TM7ab structure_element The most apparent of these changes involves the N-terminal half of TM7 (TM7ab); together with more subtle displacements in TM2 and TM3, this change in TM7ab correlates with the opening and closing of two distinct aqueous channels leading into the ion-binding sites from the extracellular solution. DISCUSS | |
| 123 126 TM2 structure_element The most apparent of these changes involves the N-terminal half of TM7 (TM7ab); together with more subtle displacements in TM2 and TM3, this change in TM7ab correlates with the opening and closing of two distinct aqueous channels leading into the ion-binding sites from the extracellular solution. DISCUSS | |
| 131 134 TM3 structure_element The most apparent of these changes involves the N-terminal half of TM7 (TM7ab); together with more subtle displacements in TM2 and TM3, this change in TM7ab correlates with the opening and closing of two distinct aqueous channels leading into the ion-binding sites from the extracellular solution. DISCUSS | |
| 151 156 TM7ab structure_element The most apparent of these changes involves the N-terminal half of TM7 (TM7ab); together with more subtle displacements in TM2 and TM3, this change in TM7ab correlates with the opening and closing of two distinct aqueous channels leading into the ion-binding sites from the extracellular solution. DISCUSS | |
| 213 229 aqueous channels site The most apparent of these changes involves the N-terminal half of TM7 (TM7ab); together with more subtle displacements in TM2 and TM3, this change in TM7ab correlates with the opening and closing of two distinct aqueous channels leading into the ion-binding sites from the extracellular solution. DISCUSS | |
| 247 264 ion-binding sites site The most apparent of these changes involves the N-terminal half of TM7 (TM7ab); together with more subtle displacements in TM2 and TM3, this change in TM7ab correlates with the opening and closing of two distinct aqueous channels leading into the ion-binding sites from the extracellular solution. DISCUSS | |
| 30 33 TM7 structure_element Interestingly, the bending of TM7 associated with the occlusion of the ion-binding sites also unlocks its interaction with TM6, and thus enables TM6 and TM1 to freely slide to the inward-facing conformation. DISCUSS | |
| 71 88 ion-binding sites site Interestingly, the bending of TM7 associated with the occlusion of the ion-binding sites also unlocks its interaction with TM6, and thus enables TM6 and TM1 to freely slide to the inward-facing conformation. DISCUSS | |
| 123 126 TM6 structure_element Interestingly, the bending of TM7 associated with the occlusion of the ion-binding sites also unlocks its interaction with TM6, and thus enables TM6 and TM1 to freely slide to the inward-facing conformation. DISCUSS | |
| 145 148 TM6 structure_element Interestingly, the bending of TM7 associated with the occlusion of the ion-binding sites also unlocks its interaction with TM6, and thus enables TM6 and TM1 to freely slide to the inward-facing conformation. DISCUSS | |
| 153 156 TM1 structure_element Interestingly, the bending of TM7 associated with the occlusion of the ion-binding sites also unlocks its interaction with TM6, and thus enables TM6 and TM1 to freely slide to the inward-facing conformation. DISCUSS | |
| 180 193 inward-facing protein_state Interestingly, the bending of TM7 associated with the occlusion of the ion-binding sites also unlocks its interaction with TM6, and thus enables TM6 and TM1 to freely slide to the inward-facing conformation. DISCUSS | |
| 4 22 crystal structures evidence The crystal structures of NCX_Mj reported here, with either Na+, Ca2+, Sr2+ or H+ bound, capture the exchanger in different conformational states. DISCUSS | |
| 26 32 NCX_Mj protein The crystal structures of NCX_Mj reported here, with either Na+, Ca2+, Sr2+ or H+ bound, capture the exchanger in different conformational states. DISCUSS | |
| 60 64 Na+, chemical The crystal structures of NCX_Mj reported here, with either Na+, Ca2+, Sr2+ or H+ bound, capture the exchanger in different conformational states. DISCUSS | |
| 65 70 Ca2+, chemical The crystal structures of NCX_Mj reported here, with either Na+, Ca2+, Sr2+ or H+ bound, capture the exchanger in different conformational states. DISCUSS | |
| 71 75 Sr2+ chemical The crystal structures of NCX_Mj reported here, with either Na+, Ca2+, Sr2+ or H+ bound, capture the exchanger in different conformational states. DISCUSS | |
| 79 81 H+ chemical The crystal structures of NCX_Mj reported here, with either Na+, Ca2+, Sr2+ or H+ bound, capture the exchanger in different conformational states. DISCUSS | |
| 82 87 bound protein_state The crystal structures of NCX_Mj reported here, with either Na+, Ca2+, Sr2+ or H+ bound, capture the exchanger in different conformational states. DISCUSS | |
| 101 110 exchanger protein_type The crystal structures of NCX_Mj reported here, with either Na+, Ca2+, Sr2+ or H+ bound, capture the exchanger in different conformational states. DISCUSS | |
| 115 126 transporter protein_type These states can only represent a subset among all possible, but they ought to reflect inherent preferences of the transporter, modulated by the experimental conditions. DISCUSS | |
| 20 27 crystal evidence For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 31 37 NCX_Mj protein For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 41 44 LCP experimental_method For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 50 68 extracellular half structure_element For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 76 90 gating helices structure_element For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 92 95 TM6 structure_element For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 100 103 TM1 structure_element For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 190 207 ion-binding sites site For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 235 238 apo protein_state For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 240 246 low pH protein_state For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 247 256 structure evidence For example, in the crystal of NCX_Mj in LCP, the extracellular half of the gating helices (TM6 and TM1) form a lattice contact, which might ultimately restrict the degree of opening of the ion-binding sites in some cases (e.g. in the apo, low pH structure). DISCUSS | |
| 17 50 calculated free-energy landscapes evidence Nonetheless, the calculated free-energy landscapes, derived without knowledge of the experimental data, reassuringly confirm that the crystallized structures correspond to mechanistically relevant, interconverting states. DISCUSS | |
| 134 157 crystallized structures evidence Nonetheless, the calculated free-energy landscapes, derived without knowledge of the experimental data, reassuringly confirm that the crystallized structures correspond to mechanistically relevant, interconverting states. DISCUSS | |
| 4 15 simulations experimental_method The simulations also demonstrate how this landscape is drastically re-shaped upon each ion-binding event. DISCUSS | |
| 58 66 occluded protein_state Indeed, we show that it is the presence or absence of the occluded state in this landscape that explains the antiport function of NCX_Mj and its 3Na+:1Ca2+ stoichiometry. DISCUSS | |
| 130 136 NCX_Mj protein Indeed, we show that it is the presence or absence of the occluded state in this landscape that explains the antiport function of NCX_Mj and its 3Na+:1Ca2+ stoichiometry. DISCUSS | |
| 146 149 Na+ chemical Indeed, we show that it is the presence or absence of the occluded state in this landscape that explains the antiport function of NCX_Mj and its 3Na+:1Ca2+ stoichiometry. DISCUSS | |
| 151 155 Ca2+ chemical Indeed, we show that it is the presence or absence of the occluded state in this landscape that explains the antiport function of NCX_Mj and its 3Na+:1Ca2+ stoichiometry. DISCUSS | |
| 89 101 transporters protein_type We posit that a similar principle might govern the alternating-access mechanism in other transporters; that is, we anticipate that for both symporters and antiporters, it is the feasibility of the occluded state, encoded in the protein conformational free-energy landscape and its dependence on substrate binding, that ultimately explains their specific coupling mechanisms. DISCUSS | |
| 140 150 symporters protein_type We posit that a similar principle might govern the alternating-access mechanism in other transporters; that is, we anticipate that for both symporters and antiporters, it is the feasibility of the occluded state, encoded in the protein conformational free-energy landscape and its dependence on substrate binding, that ultimately explains their specific coupling mechanisms. DISCUSS | |
| 155 166 antiporters protein_type We posit that a similar principle might govern the alternating-access mechanism in other transporters; that is, we anticipate that for both symporters and antiporters, it is the feasibility of the occluded state, encoded in the protein conformational free-energy landscape and its dependence on substrate binding, that ultimately explains their specific coupling mechanisms. DISCUSS | |
| 197 205 occluded protein_state We posit that a similar principle might govern the alternating-access mechanism in other transporters; that is, we anticipate that for both symporters and antiporters, it is the feasibility of the occluded state, encoded in the protein conformational free-energy landscape and its dependence on substrate binding, that ultimately explains their specific coupling mechanisms. DISCUSS | |
| 228 272 protein conformational free-energy landscape evidence We posit that a similar principle might govern the alternating-access mechanism in other transporters; that is, we anticipate that for both symporters and antiporters, it is the feasibility of the occluded state, encoded in the protein conformational free-energy landscape and its dependence on substrate binding, that ultimately explains their specific coupling mechanisms. DISCUSS | |
| 122 128 NCX_Mj protein In multiple ways, our findings provide an explanation for, existing functional, biochemical and biophysical data for both NCX_Mj and its eukaryotic homologues. DISCUSS | |
| 137 147 eukaryotic taxonomy_domain In multiple ways, our findings provide an explanation for, existing functional, biochemical and biophysical data for both NCX_Mj and its eukaryotic homologues. DISCUSS | |
| 48 70 ion-binding affinities evidence The striking quantitative agreement between the ion-binding affinities inferred from our crystallographic titrations and the Km and K1/2 values previously deduced from functional assays has been discussed above. DISCUSS | |
| 89 116 crystallographic titrations experimental_method The striking quantitative agreement between the ion-binding affinities inferred from our crystallographic titrations and the Km and K1/2 values previously deduced from functional assays has been discussed above. DISCUSS | |
| 125 127 Km evidence The striking quantitative agreement between the ion-binding affinities inferred from our crystallographic titrations and the Km and K1/2 values previously deduced from functional assays has been discussed above. DISCUSS | |
| 132 143 K1/2 values evidence The striking quantitative agreement between the ion-binding affinities inferred from our crystallographic titrations and the Km and K1/2 values previously deduced from functional assays has been discussed above. DISCUSS | |
| 168 185 functional assays experimental_method The striking quantitative agreement between the ion-binding affinities inferred from our crystallographic titrations and the Km and K1/2 values previously deduced from functional assays has been discussed above. DISCUSS | |
| 113 119 NCX_Mj protein Consistent with that finding, mutations that have been shown to inactivate or diminish the transport activity of NCX_Mj and cardiac NCX perfectly map to the first ion-coordination shell in our NCX_Mj structures (Supplementary Fig. 4c-d). DISCUSS | |
| 132 135 NCX protein_type Consistent with that finding, mutations that have been shown to inactivate or diminish the transport activity of NCX_Mj and cardiac NCX perfectly map to the first ion-coordination shell in our NCX_Mj structures (Supplementary Fig. 4c-d). DISCUSS | |
| 193 199 NCX_Mj protein Consistent with that finding, mutations that have been shown to inactivate or diminish the transport activity of NCX_Mj and cardiac NCX perfectly map to the first ion-coordination shell in our NCX_Mj structures (Supplementary Fig. 4c-d). DISCUSS | |
| 200 210 structures evidence Consistent with that finding, mutations that have been shown to inactivate or diminish the transport activity of NCX_Mj and cardiac NCX perfectly map to the first ion-coordination shell in our NCX_Mj structures (Supplementary Fig. 4c-d). DISCUSS | |
| 4 25 crystallographic data evidence The crystallographic data also provides the long-sought structural basis for the ‘two-site’ model proposed to describe competitive cation binding in eukaryotic NCX, underscoring the relevance of these studies of NCX_Mj as a prototypical Na+/Ca2+ exchanger. DISCUSS | |
| 149 159 eukaryotic taxonomy_domain The crystallographic data also provides the long-sought structural basis for the ‘two-site’ model proposed to describe competitive cation binding in eukaryotic NCX, underscoring the relevance of these studies of NCX_Mj as a prototypical Na+/Ca2+ exchanger. DISCUSS | |
| 160 163 NCX protein_type The crystallographic data also provides the long-sought structural basis for the ‘two-site’ model proposed to describe competitive cation binding in eukaryotic NCX, underscoring the relevance of these studies of NCX_Mj as a prototypical Na+/Ca2+ exchanger. DISCUSS | |
| 212 218 NCX_Mj protein The crystallographic data also provides the long-sought structural basis for the ‘two-site’ model proposed to describe competitive cation binding in eukaryotic NCX, underscoring the relevance of these studies of NCX_Mj as a prototypical Na+/Ca2+ exchanger. DISCUSS | |
| 237 255 Na+/Ca2+ exchanger protein_type The crystallographic data also provides the long-sought structural basis for the ‘two-site’ model proposed to describe competitive cation binding in eukaryotic NCX, underscoring the relevance of these studies of NCX_Mj as a prototypical Na+/Ca2+ exchanger. DISCUSS | |
| 18 36 crystal titrations experimental_method Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 66 69 Na+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 70 74 Ca2+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 91 95 Sint site Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 100 103 SCa site Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 111 115 Ca2+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 120 123 Na+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 162 165 Na+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 211 214 Na+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 239 242 Na+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 261 266 water chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 280 284 Smid site Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 313 317 Ca2+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 334 350 Hill coefficient evidence Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 361 364 Na+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 389 393 Ca2+ chemical Specifically, our crystal titrations suggest that, during forward Na+/Ca2+ exchange, sites Sint and SCa, which Ca2+ and Na+ compete for, can be grouped into one; Na+ binding to these sites does not require high Na+ concentrations, and two Na+ ions along with a water molecule (at Smid) are sufficient to displace Ca2+, explaining the Hill coefficient of ~2 for Na+-dependent inhibition of Ca2+ fluxes. DISCUSS | |
| 4 8 Sext site The Sext site, by contrast, might be thought as an activation site for inward Na+ translocation, since this is where the third Na+ ion binds at high Na+ concentration, enabling the transition to the occluded state. DISCUSS | |
| 51 66 activation site site The Sext site, by contrast, might be thought as an activation site for inward Na+ translocation, since this is where the third Na+ ion binds at high Na+ concentration, enabling the transition to the occluded state. DISCUSS | |
| 78 81 Na+ chemical The Sext site, by contrast, might be thought as an activation site for inward Na+ translocation, since this is where the third Na+ ion binds at high Na+ concentration, enabling the transition to the occluded state. DISCUSS | |
| 127 130 Na+ chemical The Sext site, by contrast, might be thought as an activation site for inward Na+ translocation, since this is where the third Na+ ion binds at high Na+ concentration, enabling the transition to the occluded state. DISCUSS | |
| 149 152 Na+ chemical The Sext site, by contrast, might be thought as an activation site for inward Na+ translocation, since this is where the third Na+ ion binds at high Na+ concentration, enabling the transition to the occluded state. DISCUSS | |
| 199 207 occluded protein_state The Sext site, by contrast, might be thought as an activation site for inward Na+ translocation, since this is where the third Na+ ion binds at high Na+ concentration, enabling the transition to the occluded state. DISCUSS | |
| 26 30 Ca2+ chemical Interestingly, binding of Ca2+ to Smid appears to be also possible, but available evidence indicates that this event transiently blocks the exchange cycle. DISCUSS | |
| 34 38 Smid site Interestingly, binding of Ca2+ to Smid appears to be also possible, but available evidence indicates that this event transiently blocks the exchange cycle. DISCUSS | |
| 8 18 structures evidence Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 22 28 NCX_Mj protein Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 29 37 bound to protein_state Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 38 42 Cd2+ chemical Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 46 50 Mn2+ chemical Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 104 108 Smid site Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 123 127 Sr2+ chemical Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 142 145 SCa site Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 182 185 NCX protein_type Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 199 206 calcium chemical Indeed, structures of NCX_Mj bound to Cd2+ or Mn2+, both of which inhibit transport, show these ions at Smid; by contrast, Sr2+ binds only to SCa, and accordingly, is transported by NCX similarly to calcium. DISCUSS | |
| 37 43 NCX_Mj protein Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 72 76 Ca2+ chemical Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 80 83 Na+ chemical Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 147 174 hydrogen-deuterium exchange experimental_method Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 176 179 HDX experimental_method Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 184 190 NCX_Mj protein Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 199 207 presence protein_state Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 211 221 absence of protein_state Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 259 273 outward-facing protein_state Lastly, our theory that occlusion of NCX_Mj is selectively induced upon Ca2+ or Na+ recognition is consonant with a recent analysis of the rate of hydrogen-deuterium exchange (HDX) in NCX_Mj, in the presence or absence of these ions, in conditions that favor outward-facing conformations. DISCUSS | |
| 36 40 Ca2+ chemical Specifically, saturating amounts of Ca2+ or Na+ resulted in a noticeable slowdown in the HDX rate for extracellular portions of the α-repeat helices. DISCUSS | |
| 44 47 Na+ chemical Specifically, saturating amounts of Ca2+ or Na+ resulted in a noticeable slowdown in the HDX rate for extracellular portions of the α-repeat helices. DISCUSS | |
| 89 97 HDX rate evidence Specifically, saturating amounts of Ca2+ or Na+ resulted in a noticeable slowdown in the HDX rate for extracellular portions of the α-repeat helices. DISCUSS | |
| 132 148 α-repeat helices structure_element Specifically, saturating amounts of Ca2+ or Na+ resulted in a noticeable slowdown in the HDX rate for extracellular portions of the α-repeat helices. DISCUSS | |
| 231 248 ion-binding sites site We interpret these observations as reflecting that the solvent accessibility of the protein interior is diminished upon ion recognition, consistent with our finding that opening and closing of extracellular aqueous pathways to the ion-binding sites depend on ion occupancy state. DISCUSS | |
| 80 88 occluded protein_state In addition, the increased compactness of the protein tertiary structure in the occluded state would also slow down the dynamics of the secondary-structure elements, and thus further reduce the HDX rate. DISCUSS | |
| 194 202 HDX rate evidence In addition, the increased compactness of the protein tertiary structure in the occluded state would also slow down the dynamics of the secondary-structure elements, and thus further reduce the HDX rate. DISCUSS | |
| 70 78 HDX rate evidence Our data would also explain the observation that the reduction in the HDX rate is comparable for Na+ and Ca2+, as well as the finding that the degree of deuterium incorporation remains non-negligible even under saturating ion concentrations. DISCUSS | |
| 97 100 Na+ chemical Our data would also explain the observation that the reduction in the HDX rate is comparable for Na+ and Ca2+, as well as the finding that the degree of deuterium incorporation remains non-negligible even under saturating ion concentrations. DISCUSS | |
| 105 109 Ca2+ chemical Our data would also explain the observation that the reduction in the HDX rate is comparable for Na+ and Ca2+, as well as the finding that the degree of deuterium incorporation remains non-negligible even under saturating ion concentrations. DISCUSS | |
| 7 40 calculated free-energy landscapes evidence As the calculated free-energy landscapes show, Na+ and Ca2+ induce the occlusion of the transporter in a comparable manner, and yet the ion-bound states retain the ability to explore conformations that are partially or fully open to the extracellular solution, precisely so as to be able to unload and re-load the substrates. DISCUSS | |
| 47 50 Na+ chemical As the calculated free-energy landscapes show, Na+ and Ca2+ induce the occlusion of the transporter in a comparable manner, and yet the ion-bound states retain the ability to explore conformations that are partially or fully open to the extracellular solution, precisely so as to be able to unload and re-load the substrates. DISCUSS | |
| 55 59 Ca2+ chemical As the calculated free-energy landscapes show, Na+ and Ca2+ induce the occlusion of the transporter in a comparable manner, and yet the ion-bound states retain the ability to explore conformations that are partially or fully open to the extracellular solution, precisely so as to be able to unload and re-load the substrates. DISCUSS | |
| 88 99 transporter protein_type As the calculated free-energy landscapes show, Na+ and Ca2+ induce the occlusion of the transporter in a comparable manner, and yet the ion-bound states retain the ability to explore conformations that are partially or fully open to the extracellular solution, precisely so as to be able to unload and re-load the substrates. DISCUSS | |
| 136 145 ion-bound protein_state As the calculated free-energy landscapes show, Na+ and Ca2+ induce the occlusion of the transporter in a comparable manner, and yet the ion-bound states retain the ability to explore conformations that are partially or fully open to the extracellular solution, precisely so as to be able to unload and re-load the substrates. DISCUSS | |
| 219 229 fully open protein_state As the calculated free-energy landscapes show, Na+ and Ca2+ induce the occlusion of the transporter in a comparable manner, and yet the ion-bound states retain the ability to explore conformations that are partially or fully open to the extracellular solution, precisely so as to be able to unload and re-load the substrates. DISCUSS | |
| 0 3 Na+ chemical Na+ binding to outward-facing NCX_Mj. FIG | |
| 15 29 outward-facing protein_state Na+ binding to outward-facing NCX_Mj. FIG | |
| 30 36 NCX_Mj protein Na+ binding to outward-facing NCX_Mj. FIG | |
| 12 21 structure evidence (a) Overall structure of native outward-facing NCX_Mj from crystals grown in 150 mM Na+. FIG | |
| 25 31 native protein_state (a) Overall structure of native outward-facing NCX_Mj from crystals grown in 150 mM Na+. FIG | |
| 32 46 outward-facing protein_state (a) Overall structure of native outward-facing NCX_Mj from crystals grown in 150 mM Na+. FIG | |
| 47 53 NCX_Mj protein (a) Overall structure of native outward-facing NCX_Mj from crystals grown in 150 mM Na+. FIG | |
| 59 73 crystals grown experimental_method (a) Overall structure of native outward-facing NCX_Mj from crystals grown in 150 mM Na+. FIG | |
| 84 87 Na+ chemical (a) Overall structure of native outward-facing NCX_Mj from crystals grown in 150 mM Na+. FIG | |
| 36 39 Na+ chemical Colored spheres represent the bound Na+ (green) and water (red). FIG | |
| 52 57 water chemical Colored spheres represent the bound Na+ (green) and water (red). FIG | |
| 50 71 central binding sites site (b) Structural details and definition of the four central binding sites. FIG | |
| 4 20 electron density evidence The electron density (grey mesh, 1.9 Å Fo-Fc ion omit map contoured at 4σ) at Smid was modeled as water (red sphere) and those at Sext, SCa and Sint as Na+ ions (green spheres). FIG | |
| 39 57 Fo-Fc ion omit map evidence The electron density (grey mesh, 1.9 Å Fo-Fc ion omit map contoured at 4σ) at Smid was modeled as water (red sphere) and those at Sext, SCa and Sint as Na+ ions (green spheres). FIG | |
| 78 82 Smid site The electron density (grey mesh, 1.9 Å Fo-Fc ion omit map contoured at 4σ) at Smid was modeled as water (red sphere) and those at Sext, SCa and Sint as Na+ ions (green spheres). FIG | |
| 98 103 water chemical The electron density (grey mesh, 1.9 Å Fo-Fc ion omit map contoured at 4σ) at Smid was modeled as water (red sphere) and those at Sext, SCa and Sint as Na+ ions (green spheres). FIG | |
| 130 134 Sext site The electron density (grey mesh, 1.9 Å Fo-Fc ion omit map contoured at 4σ) at Smid was modeled as water (red sphere) and those at Sext, SCa and Sint as Na+ ions (green spheres). FIG | |
| 136 139 SCa site The electron density (grey mesh, 1.9 Å Fo-Fc ion omit map contoured at 4σ) at Smid was modeled as water (red sphere) and those at Sext, SCa and Sint as Na+ ions (green spheres). FIG | |
| 144 148 Sint site The electron density (grey mesh, 1.9 Å Fo-Fc ion omit map contoured at 4σ) at Smid was modeled as water (red sphere) and those at Sext, SCa and Sint as Na+ ions (green spheres). FIG | |
| 152 155 Na+ chemical The electron density (grey mesh, 1.9 Å Fo-Fc ion omit map contoured at 4σ) at Smid was modeled as water (red sphere) and those at Sext, SCa and Sint as Na+ ions (green spheres). FIG | |
| 89 92 Na+ chemical Further details are shown in Supplementary Fig. 1. (c) Concentration-dependent change in Na+ occupancy (see also Table 1). FIG | |
| 4 25 Fo – Fc ion-omit maps evidence All Fo – Fc ion-omit maps are calculated to 2.4 Å and contoured at 3σ for comparison. FIG | |
| 20 24 A206 residue_name_number The displacement of A206 reflects the [Na+]-dependent conformational change from the partially open to the occluded state (observed at low and high Na+ concentrations, respectively). FIG | |
| 39 42 Na+ chemical The displacement of A206 reflects the [Na+]-dependent conformational change from the partially open to the occluded state (observed at low and high Na+ concentrations, respectively). FIG | |
| 85 99 partially open protein_state The displacement of A206 reflects the [Na+]-dependent conformational change from the partially open to the occluded state (observed at low and high Na+ concentrations, respectively). FIG | |
| 107 115 occluded protein_state The displacement of A206 reflects the [Na+]-dependent conformational change from the partially open to the occluded state (observed at low and high Na+ concentrations, respectively). FIG | |
| 148 151 Na+ chemical The displacement of A206 reflects the [Na+]-dependent conformational change from the partially open to the occluded state (observed at low and high Na+ concentrations, respectively). FIG | |
| 9 12 Na+ chemical At 20 mM Na+, both conformations co-exist. FIG | |
| 75 80 water chemical No significant changes were observed in the side-chains involved in ion or water coordination at the SCa, Sint and Smid sites. FIG | |
| 101 104 SCa site No significant changes were observed in the side-chains involved in ion or water coordination at the SCa, Sint and Smid sites. FIG | |
| 106 110 Sint site No significant changes were observed in the side-chains involved in ion or water coordination at the SCa, Sint and Smid sites. FIG | |
| 115 119 Smid site No significant changes were observed in the side-chains involved in ion or water coordination at the SCa, Sint and Smid sites. FIG | |
| 0 3 Na+ chemical Na+-occupancy dependent conformational change in NCX_Mj. FIG | |
| 49 55 NCX_Mj protein Na+-occupancy dependent conformational change in NCX_Mj. FIG | |
| 4 19 Superimposition experimental_method (a) Superimposition of the NCX_Mj crystal structures obtained in high (100 mM, cyan cylinders) and low (10 mM, brown cylinders) Na+ concentrations. FIG | |
| 27 33 NCX_Mj protein (a) Superimposition of the NCX_Mj crystal structures obtained in high (100 mM, cyan cylinders) and low (10 mM, brown cylinders) Na+ concentrations. FIG | |
| 34 52 crystal structures evidence (a) Superimposition of the NCX_Mj crystal structures obtained in high (100 mM, cyan cylinders) and low (10 mM, brown cylinders) Na+ concentrations. FIG | |
| 128 131 Na+ chemical (a) Superimposition of the NCX_Mj crystal structures obtained in high (100 mM, cyan cylinders) and low (10 mM, brown cylinders) Na+ concentrations. FIG | |
| 25 34 interface site (b) Close-up view of the interface between TM6 and TM7ab in the NCX_Mj structures obtained at high and low Na+ concentrations (top and lower panels, respectively). FIG | |
| 43 46 TM6 structure_element (b) Close-up view of the interface between TM6 and TM7ab in the NCX_Mj structures obtained at high and low Na+ concentrations (top and lower panels, respectively). FIG | |
| 51 56 TM7ab structure_element (b) Close-up view of the interface between TM6 and TM7ab in the NCX_Mj structures obtained at high and low Na+ concentrations (top and lower panels, respectively). FIG | |
| 64 70 NCX_Mj protein (b) Close-up view of the interface between TM6 and TM7ab in the NCX_Mj structures obtained at high and low Na+ concentrations (top and lower panels, respectively). FIG | |
| 71 81 structures evidence (b) Close-up view of the interface between TM6 and TM7ab in the NCX_Mj structures obtained at high and low Na+ concentrations (top and lower panels, respectively). FIG | |
| 107 110 Na+ chemical (b) Close-up view of the interface between TM6 and TM7ab in the NCX_Mj structures obtained at high and low Na+ concentrations (top and lower panels, respectively). FIG | |
| 47 56 structure evidence Residues forming van-der-Waals contacts in the structure at low Na+ concentration are shown in detail. FIG | |
| 60 63 low protein_state Residues forming van-der-Waals contacts in the structure at low Na+ concentration are shown in detail. FIG | |
| 64 67 Na+ chemical Residues forming van-der-Waals contacts in the structure at low Na+ concentration are shown in detail. FIG | |
| 25 42 Na+-binding sites site (c) Close-up view of the Na+-binding sites. FIG | |
| 11 15 Sext site The vacant Sext site in the structure at low Na+ concentration is indicated with a white sphere. FIG | |
| 28 37 structure evidence The vacant Sext site in the structure at low Na+ concentration is indicated with a white sphere. FIG | |
| 41 44 low protein_state The vacant Sext site in the structure at low Na+ concentration is indicated with a white sphere. FIG | |
| 45 48 Na+ chemical The vacant Sext site in the structure at low Na+ concentration is indicated with a white sphere. FIG | |
| 56 60 A206 residue_name_number Residues surrounding this site are also indicated; note A206 (labeled in red) coordinates Na+ at Sext via its backbone carbonyl oxygen. FIG | |
| 78 89 coordinates bond_interaction Residues surrounding this site are also indicated; note A206 (labeled in red) coordinates Na+ at Sext via its backbone carbonyl oxygen. FIG | |
| 90 93 Na+ chemical Residues surrounding this site are also indicated; note A206 (labeled in red) coordinates Na+ at Sext via its backbone carbonyl oxygen. FIG | |
| 97 101 Sext site Residues surrounding this site are also indicated; note A206 (labeled in red) coordinates Na+ at Sext via its backbone carbonyl oxygen. FIG | |
| 47 64 ion binding sites site (d) Extracellular solvent accessibility of the ion binding sites in the structures at high and low [Na+]. FIG | |
| 72 82 structures evidence (d) Extracellular solvent accessibility of the ion binding sites in the structures at high and low [Na+]. FIG | |
| 86 90 high protein_state (d) Extracellular solvent accessibility of the ion binding sites in the structures at high and low [Na+]. FIG | |
| 95 98 low protein_state (d) Extracellular solvent accessibility of the ion binding sites in the structures at high and low [Na+]. FIG | |
| 100 103 Na+ chemical (d) Extracellular solvent accessibility of the ion binding sites in the structures at high and low [Na+]. FIG | |
| 9 25 solvent channels site Putative solvent channels are represented as light-purple surfaces. FIG | |
| 28 31 apo protein_state Divalent cation binding and apo structure of NCX_Mj. (a) A single Sr2+ (dark blue sphere) binds at SCa in crystals titrated with 10 mM Sr2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 32 41 structure evidence Divalent cation binding and apo structure of NCX_Mj. (a) A single Sr2+ (dark blue sphere) binds at SCa in crystals titrated with 10 mM Sr2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 45 51 NCX_Mj protein Divalent cation binding and apo structure of NCX_Mj. (a) A single Sr2+ (dark blue sphere) binds at SCa in crystals titrated with 10 mM Sr2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 66 70 Sr2+ chemical Divalent cation binding and apo structure of NCX_Mj. (a) A single Sr2+ (dark blue sphere) binds at SCa in crystals titrated with 10 mM Sr2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 99 102 SCa site Divalent cation binding and apo structure of NCX_Mj. (a) A single Sr2+ (dark blue sphere) binds at SCa in crystals titrated with 10 mM Sr2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 106 123 crystals titrated experimental_method Divalent cation binding and apo structure of NCX_Mj. (a) A single Sr2+ (dark blue sphere) binds at SCa in crystals titrated with 10 mM Sr2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 135 139 Sr2+ chemical Divalent cation binding and apo structure of NCX_Mj. (a) A single Sr2+ (dark blue sphere) binds at SCa in crystals titrated with 10 mM Sr2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 151 154 Na+ chemical Divalent cation binding and apo structure of NCX_Mj. (a) A single Sr2+ (dark blue sphere) binds at SCa in crystals titrated with 10 mM Sr2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 21 25 Sr2+ chemical Residues involved in Sr2+ coordination are labeled. FIG | |
| 98 107 Na+-bound protein_state There are no significant changes in the side-chains involved in ion coordination, relative to the Na+-bound state. FIG | |
| 0 3 T50 residue_name_number T50 and T209 (labeled in red) coordinate Sr2+ through their backbone carbonyl-oxygen atoms. FIG | |
| 8 12 T209 residue_name_number T50 and T209 (labeled in red) coordinate Sr2+ through their backbone carbonyl-oxygen atoms. FIG | |
| 30 40 coordinate bond_interaction T50 and T209 (labeled in red) coordinate Sr2+ through their backbone carbonyl-oxygen atoms. FIG | |
| 41 45 Sr2+ chemical T50 and T209 (labeled in red) coordinate Sr2+ through their backbone carbonyl-oxygen atoms. FIG | |
| 5 8 Na+ chemical High Na+ concentration (100 mM) completely displaces Sr2+ and reverts NCX_Mj to the occluded state (right panel). FIG | |
| 53 57 Sr2+ chemical High Na+ concentration (100 mM) completely displaces Sr2+ and reverts NCX_Mj to the occluded state (right panel). FIG | |
| 70 76 NCX_Mj protein High Na+ concentration (100 mM) completely displaces Sr2+ and reverts NCX_Mj to the occluded state (right panel). FIG | |
| 84 92 occluded protein_state High Na+ concentration (100 mM) completely displaces Sr2+ and reverts NCX_Mj to the occluded state (right panel). FIG | |
| 25 41 Fo – Fc omit map evidence The contour level of the Fo – Fc omit map of the structure at high Na+ concentration was lowered (to 4σ) so as to visualize the density from Na+ ions and H2O. FIG | |
| 49 58 structure evidence The contour level of the Fo – Fc omit map of the structure at high Na+ concentration was lowered (to 4σ) so as to visualize the density from Na+ ions and H2O. FIG | |
| 67 70 Na+ chemical The contour level of the Fo – Fc omit map of the structure at high Na+ concentration was lowered (to 4σ) so as to visualize the density from Na+ ions and H2O. FIG | |
| 128 135 density evidence The contour level of the Fo – Fc omit map of the structure at high Na+ concentration was lowered (to 4σ) so as to visualize the density from Na+ ions and H2O. FIG | |
| 141 144 Na+ chemical The contour level of the Fo – Fc omit map of the structure at high Na+ concentration was lowered (to 4σ) so as to visualize the density from Na+ ions and H2O. FIG | |
| 154 157 H2O chemical The contour level of the Fo – Fc omit map of the structure at high Na+ concentration was lowered (to 4σ) so as to visualize the density from Na+ ions and H2O. FIG | |
| 4 8 Ca2+ chemical (b) Ca2+ (tanned spheres) binds either to SCa or Smid in crystals titrated with 10 mM Ca2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 42 45 SCa site (b) Ca2+ (tanned spheres) binds either to SCa or Smid in crystals titrated with 10 mM Ca2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 49 53 Smid site (b) Ca2+ (tanned spheres) binds either to SCa or Smid in crystals titrated with 10 mM Ca2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 57 74 crystals titrated experimental_method (b) Ca2+ (tanned spheres) binds either to SCa or Smid in crystals titrated with 10 mM Ca2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 86 90 Ca2+ chemical (b) Ca2+ (tanned spheres) binds either to SCa or Smid in crystals titrated with 10 mM Ca2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 102 105 Na+ chemical (b) Ca2+ (tanned spheres) binds either to SCa or Smid in crystals titrated with 10 mM Ca2+ and 2.5 mM Na+ (see also Supplementary Fig. 2). FIG | |
| 60 75 Superimposition experimental_method The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 79 85 NCX_Mj protein The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 86 96 structures evidence The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 113 116 Na+ chemical The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 169 179 absence of protein_state The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 180 183 Na+ chemical The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 188 192 pH 4 protein_state The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 223 226 apo protein_state The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 259 276 ion-binding sites site The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 284 287 apo protein_state The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 292 299 high H+ protein_state The relative occupancies are 55% and 45%, respectively. (c) Superimposition of NCX_Mj structures obtained at low Na+ concentration (10 mM) and pH 6.5 (brown) and in the absence of Na+ and pH 4 (light green), referred to as apo state. (d) Close-up view of the ion-binding sites in the apo (or high H+) state. FIG | |
| 19 22 E54 residue_name_number The side chains of E54 and E213 from the low Na+ structure are also shown (light brown) for comparison. FIG | |
| 27 31 E213 residue_name_number The side chains of E54 and E213 from the low Na+ structure are also shown (light brown) for comparison. FIG | |
| 41 48 low Na+ protein_state The side chains of E54 and E213 from the low Na+ structure are also shown (light brown) for comparison. FIG | |
| 49 58 structure evidence The side chains of E54 and E213 from the low Na+ structure are also shown (light brown) for comparison. FIG | |
| 36 40 Sint site White spheres indicate the location Sint, Smid SCa. (e) Extracellular solvent accessibility of the ion-binding sites in apo NCX_Mj. FIG | |
| 42 46 Smid site White spheres indicate the location Sint, Smid SCa. (e) Extracellular solvent accessibility of the ion-binding sites in apo NCX_Mj. FIG | |
| 47 50 SCa site White spheres indicate the location Sint, Smid SCa. (e) Extracellular solvent accessibility of the ion-binding sites in apo NCX_Mj. FIG | |
| 99 116 ion-binding sites site White spheres indicate the location Sint, Smid SCa. (e) Extracellular solvent accessibility of the ion-binding sites in apo NCX_Mj. FIG | |
| 120 123 apo protein_state White spheres indicate the location Sint, Smid SCa. (e) Extracellular solvent accessibility of the ion-binding sites in apo NCX_Mj. FIG | |
| 124 130 NCX_Mj protein White spheres indicate the location Sint, Smid SCa. (e) Extracellular solvent accessibility of the ion-binding sites in apo NCX_Mj. FIG | |
| 27 36 structure evidence Spontaneous changes in the structure of outward-occluded, fully Na+-occupied NCX_Mj (PDB code 3V5U) upon sequential displacement of Na+. FIG | |
| 40 56 outward-occluded protein_state Spontaneous changes in the structure of outward-occluded, fully Na+-occupied NCX_Mj (PDB code 3V5U) upon sequential displacement of Na+. FIG | |
| 58 76 fully Na+-occupied protein_state Spontaneous changes in the structure of outward-occluded, fully Na+-occupied NCX_Mj (PDB code 3V5U) upon sequential displacement of Na+. FIG | |
| 77 83 NCX_Mj protein Spontaneous changes in the structure of outward-occluded, fully Na+-occupied NCX_Mj (PDB code 3V5U) upon sequential displacement of Na+. FIG | |
| 132 135 Na+ chemical Spontaneous changes in the structure of outward-occluded, fully Na+-occupied NCX_Mj (PDB code 3V5U) upon sequential displacement of Na+. FIG | |
| 19 29 simulation experimental_method (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 43 49 NCX_Mj protein (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 65 68 Na+ chemical (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 69 77 bound at protein_state (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 78 82 Sext site (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 84 87 SCa site (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 92 96 Sint site (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 139 142 Na+ chemical (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 143 156 bound only at protein_state (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 157 160 SCa site (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 165 169 Sint site (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 240 255 N-terminal half structure_element (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 259 262 TM7 structure_element (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 264 269 TM7ab structure_element (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 284 287 Na+ chemical (a) Representative simulation snapshots of NCX_Mj (Methods) with Na+ bound at Sext, SCa and Sint (orange cartoons, green spheres) and with Na+ bound only at SCa and Sint (marine cartoons, yellow spheres) (b) Close-up of the backbone of the N-terminal half of TM7 (TM7ab), in the same Na+ occupancy states depicted in (a). FIG | |
| 19 39 simulation snapshots evidence (c) Representative simulation snapshots (same as above) with Na+ bound at SCa and Sint (marine cartoons, yellow spheres) and without any Na+ bound (grey cartoons). FIG | |
| 61 64 Na+ chemical (c) Representative simulation snapshots (same as above) with Na+ bound at SCa and Sint (marine cartoons, yellow spheres) and without any Na+ bound (grey cartoons). FIG | |
| 65 73 bound at protein_state (c) Representative simulation snapshots (same as above) with Na+ bound at SCa and Sint (marine cartoons, yellow spheres) and without any Na+ bound (grey cartoons). FIG | |
| 74 77 SCa site (c) Representative simulation snapshots (same as above) with Na+ bound at SCa and Sint (marine cartoons, yellow spheres) and without any Na+ bound (grey cartoons). FIG | |
| 82 86 Sint site (c) Representative simulation snapshots (same as above) with Na+ bound at SCa and Sint (marine cartoons, yellow spheres) and without any Na+ bound (grey cartoons). FIG | |
| 125 132 without protein_state (c) Representative simulation snapshots (same as above) with Na+ bound at SCa and Sint (marine cartoons, yellow spheres) and without any Na+ bound (grey cartoons). FIG | |
| 137 140 Na+ chemical (c) Representative simulation snapshots (same as above) with Na+ bound at SCa and Sint (marine cartoons, yellow spheres) and without any Na+ bound (grey cartoons). FIG | |
| 141 146 bound protein_state (c) Representative simulation snapshots (same as above) with Na+ bound at SCa and Sint (marine cartoons, yellow spheres) and without any Na+ bound (grey cartoons). FIG | |
| 20 38 ion-binding region site (d) Close-up of the ion-binding region in the fully Na+-occupied state. FIG | |
| 46 64 fully Na+-occupied protein_state (d) Close-up of the ion-binding region in the fully Na+-occupied state. FIG | |
| 30 33 TM2 structure_element Approximate distances between TM2, TM3 and TM7 are indicated in Å. (e) Close-up of the ion-binding region in the partially Na+-occupied state. FIG | |
| 35 38 TM3 structure_element Approximate distances between TM2, TM3 and TM7 are indicated in Å. (e) Close-up of the ion-binding region in the partially Na+-occupied state. FIG | |
| 43 46 TM7 structure_element Approximate distances between TM2, TM3 and TM7 are indicated in Å. (e) Close-up of the ion-binding region in the partially Na+-occupied state. FIG | |
| 87 105 ion-binding region site Approximate distances between TM2, TM3 and TM7 are indicated in Å. (e) Close-up of the ion-binding region in the partially Na+-occupied state. FIG | |
| 113 135 partially Na+-occupied protein_state Approximate distances between TM2, TM3 and TM7 are indicated in Å. (e) Close-up of the ion-binding region in the partially Na+-occupied state. FIG | |
| 20 38 ion-binding region site (f) Close-up of the ion-binding region in the Na+-free state. (g-i) Analytical descriptors of the changes just described, calculated from the simulations of each Na+-occupancy state depicted in panels (a-f). FIG | |
| 46 54 Na+-free protein_state (f) Close-up of the ion-binding region in the Na+-free state. (g-i) Analytical descriptors of the changes just described, calculated from the simulations of each Na+-occupancy state depicted in panels (a-f). FIG | |
| 142 153 simulations experimental_method (f) Close-up of the ion-binding region in the Na+-free state. (g-i) Analytical descriptors of the changes just described, calculated from the simulations of each Na+-occupancy state depicted in panels (a-f). FIG | |
| 162 175 Na+-occupancy protein_state (f) Close-up of the ion-binding region in the Na+-free state. (g-i) Analytical descriptors of the changes just described, calculated from the simulations of each Na+-occupancy state depicted in panels (a-f). FIG | |
| 63 101 Bias-Exchange Metadynamics simulations experimental_method These descriptors were employed as collective variables in the Bias-Exchange Metadynamics simulations (Methods). FIG | |
| 4 29 Probability distributions evidence (g) Probability distributions of an analytical descriptor of the backbone hydrogen-bonding pattern in TM7ab (Eq. 2). (h) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the Sext site (Eq. 1). (i) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the SCa site (Eq. 1). FIG | |
| 74 90 hydrogen-bonding bond_interaction (g) Probability distributions of an analytical descriptor of the backbone hydrogen-bonding pattern in TM7ab (Eq. 2). (h) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the Sext site (Eq. 1). (i) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the SCa site (Eq. 1). FIG | |
| 102 107 TM7ab structure_element (g) Probability distributions of an analytical descriptor of the backbone hydrogen-bonding pattern in TM7ab (Eq. 2). (h) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the Sext site (Eq. 1). (i) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the SCa site (Eq. 1). FIG | |
| 223 227 Sext site (g) Probability distributions of an analytical descriptor of the backbone hydrogen-bonding pattern in TM7ab (Eq. 2). (h) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the Sext site (Eq. 1). (i) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the SCa site (Eq. 1). FIG | |
| 348 351 SCa site (g) Probability distributions of an analytical descriptor of the backbone hydrogen-bonding pattern in TM7ab (Eq. 2). (h) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the Sext site (Eq. 1). (i) Mean value (with standard deviation) of a quantitative descriptor of the solvent accessibility of the SCa site (Eq. 1). FIG | |
| 108 111 NCX protein_type Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated conformational free-energy landscapes for outward-facing NCX_Mj, for two different Na+-occupancy states, and for a state with no ions bound. FIG | |
| 117 165 Calculated conformational free-energy landscapes evidence Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated conformational free-energy landscapes for outward-facing NCX_Mj, for two different Na+-occupancy states, and for a state with no ions bound. FIG | |
| 170 184 outward-facing protein_state Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated conformational free-energy landscapes for outward-facing NCX_Mj, for two different Na+-occupancy states, and for a state with no ions bound. FIG | |
| 185 191 NCX_Mj protein Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated conformational free-energy landscapes for outward-facing NCX_Mj, for two different Na+-occupancy states, and for a state with no ions bound. FIG | |
| 211 214 Na+ chemical Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated conformational free-energy landscapes for outward-facing NCX_Mj, for two different Na+-occupancy states, and for a state with no ions bound. FIG | |
| 254 267 no ions bound protein_state Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated conformational free-energy landscapes for outward-facing NCX_Mj, for two different Na+-occupancy states, and for a state with no ions bound. FIG | |
| 4 15 free energy evidence The free energy is plotted as a function of two coordinates, each describing the degree of opening of the aqueous channels leading to the Sext and SCa sites, respectively (see Methods). FIG | |
| 106 122 aqueous channels site The free energy is plotted as a function of two coordinates, each describing the degree of opening of the aqueous channels leading to the Sext and SCa sites, respectively (see Methods). FIG | |
| 138 142 Sext site The free energy is plotted as a function of two coordinates, each describing the degree of opening of the aqueous channels leading to the Sext and SCa sites, respectively (see Methods). FIG | |
| 147 150 SCa site The free energy is plotted as a function of two coordinates, each describing the degree of opening of the aqueous channels leading to the Sext and SCa sites, respectively (see Methods). FIG | |
| 22 38 X-ray structures evidence Black circles map the X-ray structures of NCX_Mj obtained at high and low Na+ concentration, as well as that at low pH, reported in this study. FIG | |
| 42 48 NCX_Mj protein Black circles map the X-ray structures of NCX_Mj obtained at high and low Na+ concentration, as well as that at low pH, reported in this study. FIG | |
| 61 65 high protein_state Black circles map the X-ray structures of NCX_Mj obtained at high and low Na+ concentration, as well as that at low pH, reported in this study. FIG | |
| 70 73 low protein_state Black circles map the X-ray structures of NCX_Mj obtained at high and low Na+ concentration, as well as that at low pH, reported in this study. FIG | |
| 74 77 Na+ chemical Black circles map the X-ray structures of NCX_Mj obtained at high and low Na+ concentration, as well as that at low pH, reported in this study. FIG | |
| 112 118 low pH protein_state Black circles map the X-ray structures of NCX_Mj obtained at high and low Na+ concentration, as well as that at low pH, reported in this study. FIG | |
| 4 23 Density isosurfaces evidence (b) Density isosurfaces for water molecules within 12 Å of the ion-binding region (grey volumes), for each of the major conformational free-energy minima in each ion-occupancy state. FIG | |
| 28 33 water chemical (b) Density isosurfaces for water molecules within 12 Å of the ion-binding region (grey volumes), for each of the major conformational free-energy minima in each ion-occupancy state. FIG | |
| 63 81 ion-binding region site (b) Density isosurfaces for water molecules within 12 Å of the ion-binding region (grey volumes), for each of the major conformational free-energy minima in each ion-occupancy state. FIG | |
| 120 153 conformational free-energy minima evidence (b) Density isosurfaces for water molecules within 12 Å of the ion-binding region (grey volumes), for each of the major conformational free-energy minima in each ion-occupancy state. FIG | |
| 0 3 Na+ chemical Na+ ions are shown as green spheres. FIG | |
| 8 33 inverted-topology repeats structure_element The two inverted-topology repeats in the transporter structure (transparent cartoons) are colored differently (TM1-5, orange; TM6-10, marine). FIG | |
| 41 52 transporter protein_type The two inverted-topology repeats in the transporter structure (transparent cartoons) are colored differently (TM1-5, orange; TM6-10, marine). FIG | |
| 53 62 structure evidence The two inverted-topology repeats in the transporter structure (transparent cartoons) are colored differently (TM1-5, orange; TM6-10, marine). FIG | |
| 111 116 TM1-5 structure_element The two inverted-topology repeats in the transporter structure (transparent cartoons) are colored differently (TM1-5, orange; TM6-10, marine). FIG | |
| 126 132 TM6-10 structure_element The two inverted-topology repeats in the transporter structure (transparent cartoons) are colored differently (TM1-5, orange; TM6-10, marine). FIG | |
| 26 44 ion-binding region site (c) Close-up views of the ion-binding region in the same conformational free-energy minima. FIG | |
| 57 90 conformational free-energy minima evidence (c) Close-up views of the ion-binding region in the same conformational free-energy minima. FIG | |
| 25 28 Na+ chemical Key residues involved in Na+ and water coordination (W) are highlighted (sticks, black lines). FIG | |
| 33 38 water chemical Key residues involved in Na+ and water coordination (W) are highlighted (sticks, black lines). FIG | |
| 4 22 water-density maps evidence The water-density maps in (b) is shown here as a grey mesh. FIG | |
| 5 9 D240 residue_name_number Note D240 is protonated, while E54 and E213 are ionized. FIG | |
| 31 34 E54 residue_name_number Note D240 is protonated, while E54 and E213 are ionized. FIG | |
| 39 43 E213 residue_name_number Note D240 is protonated, while E54 and E213 are ionized. FIG | |
| 108 111 NCX protein_type Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated free-energy landscapes for outward-facing NCX_Mj, for the Ca2+ and the fully protonated state. FIG | |
| 117 150 Calculated free-energy landscapes evidence Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated free-energy landscapes for outward-facing NCX_Mj, for the Ca2+ and the fully protonated state. FIG | |
| 155 169 outward-facing protein_state Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated free-energy landscapes for outward-facing NCX_Mj, for the Ca2+ and the fully protonated state. FIG | |
| 170 176 NCX_Mj protein Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated free-energy landscapes for outward-facing NCX_Mj, for the Ca2+ and the fully protonated state. FIG | |
| 186 190 Ca2+ chemical Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated free-energy landscapes for outward-facing NCX_Mj, for the Ca2+ and the fully protonated state. FIG | |
| 199 215 fully protonated protein_state Thermodynamic basis for the proposed mechanism of substrate control of the alternating-access transition of NCX. (a) Calculated free-energy landscapes for outward-facing NCX_Mj, for the Ca2+ and the fully protonated state. FIG | |
| 4 15 free energy evidence The free energy is plotted as in Fig. 5. FIG | |
| 4 8 Ca2+ chemical For Ca2+, a map is shown in which a correction for the charge-transfer between the ion and the protein is introduced, alongside the uncorrected map (see Supplementary Notes 3-4 and Supplementary Fig. 5-6). FIG | |
| 12 15 map evidence For Ca2+, a map is shown in which a correction for the charge-transfer between the ion and the protein is introduced, alongside the uncorrected map (see Supplementary Notes 3-4 and Supplementary Fig. 5-6). FIG | |
| 144 147 map evidence For Ca2+, a map is shown in which a correction for the charge-transfer between the ion and the protein is introduced, alongside the uncorrected map (see Supplementary Notes 3-4 and Supplementary Fig. 5-6). FIG | |
| 16 19 map evidence The uncorrected map overstabilizes the open state relative to the semi-open and occluded because it also overestimates the cost of dehydration of the ion, once it is bound to the protein (this effect is negligible for Na+). FIG | |
| 39 43 open protein_state The uncorrected map overstabilizes the open state relative to the semi-open and occluded because it also overestimates the cost of dehydration of the ion, once it is bound to the protein (this effect is negligible for Na+). FIG | |
| 66 75 semi-open protein_state The uncorrected map overstabilizes the open state relative to the semi-open and occluded because it also overestimates the cost of dehydration of the ion, once it is bound to the protein (this effect is negligible for Na+). FIG | |
| 80 88 occluded protein_state The uncorrected map overstabilizes the open state relative to the semi-open and occluded because it also overestimates the cost of dehydration of the ion, once it is bound to the protein (this effect is negligible for Na+). FIG | |
| 166 174 bound to protein_state The uncorrected map overstabilizes the open state relative to the semi-open and occluded because it also overestimates the cost of dehydration of the ion, once it is bound to the protein (this effect is negligible for Na+). FIG | |
| 218 221 Na+ chemical The uncorrected map overstabilizes the open state relative to the semi-open and occluded because it also overestimates the cost of dehydration of the ion, once it is bound to the protein (this effect is negligible for Na+). FIG | |
| 22 40 crystal structures evidence Black circles map the crystal structures obtained at high Ca2+ concentration and at low pH (or high H+) reported in this study. FIG | |
| 58 62 Ca2+ chemical Black circles map the crystal structures obtained at high Ca2+ concentration and at low pH (or high H+) reported in this study. FIG | |
| 84 90 low pH protein_state Black circles map the crystal structures obtained at high Ca2+ concentration and at low pH (or high H+) reported in this study. FIG | |
| 95 102 high H+ protein_state Black circles map the crystal structures obtained at high Ca2+ concentration and at low pH (or high H+) reported in this study. FIG | |
| 4 29 Water-density isosurfaces evidence (b) Water-density isosurfaces analogous to those in Fig. 5 are shown for each of the major conformational free-energy minima in the free-energy maps. FIG | |
| 106 124 free-energy minima evidence (b) Water-density isosurfaces analogous to those in Fig. 5 are shown for each of the major conformational free-energy minima in the free-energy maps. FIG | |
| 132 148 free-energy maps evidence (b) Water-density isosurfaces analogous to those in Fig. 5 are shown for each of the major conformational free-energy minima in the free-energy maps. FIG | |
| 4 8 Ca2+ chemical The Ca2+ ion is shown as a red sphere; the protein is shown as in Fig. 5. (c) Close-up views of the ion-binding region in the same conformational free-energy minima. FIG | |
| 100 118 ion-binding region site The Ca2+ ion is shown as a red sphere; the protein is shown as in Fig. 5. (c) Close-up views of the ion-binding region in the same conformational free-energy minima. FIG | |
| 131 164 conformational free-energy minima evidence The Ca2+ ion is shown as a red sphere; the protein is shown as in Fig. 5. (c) Close-up views of the ion-binding region in the same conformational free-energy minima. FIG | |
| 25 29 Ca2+ chemical Key residues involved in Ca2+ and water coordination (W) are highlighted (sticks, black lines). FIG | |
| 34 39 water chemical Key residues involved in Ca2+ and water coordination (W) are highlighted (sticks, black lines). FIG | |
| 4 22 water-density maps evidence The water-density maps in (b) are shown here as a grey mesh. FIG | |
| 7 15 occluded protein_state In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 27 31 Ca2+ chemical In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 32 37 bound protein_state In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 39 44 helix structure_element In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 45 50 TM7ab structure_element In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 83 97 fully occupied protein_state In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 98 101 Na+ chemical In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 128 134 Ala206 residue_name_number In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 143 171 hydrogen-bonding interaction bond_interaction In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 177 183 Ser210 residue_name_number In the occluded state with Ca2+ bound, helix TM7ab bends in the same way as in the fully occupied Na+ state, as the carbonyl of Ala206 forms a hydrogen-bonding interaction with Ser210. FIG | |
| 62 65 NCX protein_type Structural mechanism of extracellular forward ion exchange in NCX. FIG | |
| 23 28 Ala47 residue_name_number The carbonyl groups of Ala47 (on TM2b) and Ala206 (on TM7b), and the side chains of Glu54 (on TM2c) and Glu213 (on TM7c) are highlighted; these are four of the key residues for ion chelation and conformational changes. FIG | |
| 33 37 TM2b structure_element The carbonyl groups of Ala47 (on TM2b) and Ala206 (on TM7b), and the side chains of Glu54 (on TM2c) and Glu213 (on TM7c) are highlighted; these are four of the key residues for ion chelation and conformational changes. FIG | |
| 43 49 Ala206 residue_name_number The carbonyl groups of Ala47 (on TM2b) and Ala206 (on TM7b), and the side chains of Glu54 (on TM2c) and Glu213 (on TM7c) are highlighted; these are four of the key residues for ion chelation and conformational changes. FIG | |
| 54 58 TM7b structure_element The carbonyl groups of Ala47 (on TM2b) and Ala206 (on TM7b), and the side chains of Glu54 (on TM2c) and Glu213 (on TM7c) are highlighted; these are four of the key residues for ion chelation and conformational changes. FIG | |
| 84 89 Glu54 residue_name_number The carbonyl groups of Ala47 (on TM2b) and Ala206 (on TM7b), and the side chains of Glu54 (on TM2c) and Glu213 (on TM7c) are highlighted; these are four of the key residues for ion chelation and conformational changes. FIG | |
| 94 98 TM2c structure_element The carbonyl groups of Ala47 (on TM2b) and Ala206 (on TM7b), and the side chains of Glu54 (on TM2c) and Glu213 (on TM7c) are highlighted; these are four of the key residues for ion chelation and conformational changes. FIG | |
| 104 110 Glu213 residue_name_number The carbonyl groups of Ala47 (on TM2b) and Ala206 (on TM7b), and the side chains of Glu54 (on TM2c) and Glu213 (on TM7c) are highlighted; these are four of the key residues for ion chelation and conformational changes. FIG | |
| 115 119 TM7c structure_element The carbonyl groups of Ala47 (on TM2b) and Ala206 (on TM7b), and the side chains of Glu54 (on TM2c) and Glu213 (on TM7c) are highlighted; these are four of the key residues for ion chelation and conformational changes. FIG | |
| 39 53 gating helices structure_element The green open cylinders represent the gating helices TM1 and TM6. FIG | |
| 54 57 TM1 structure_element The green open cylinders represent the gating helices TM1 and TM6. FIG | |
| 62 65 TM6 structure_element The green open cylinders represent the gating helices TM1 and TM6. FIG | |
| 32 50 crystal structures evidence Asterisks mark the states whose crystal structures have been determined in this study. FIG | |
| 65 98 calculated free-energy landscapes evidence These states and their connectivity can also be deduced from the calculated free-energy landscapes, which also reveal a Ca2+-loaded outward-facing occluded state, and an unloaded, fully open state. FIG | |
| 120 131 Ca2+-loaded protein_state These states and their connectivity can also be deduced from the calculated free-energy landscapes, which also reveal a Ca2+-loaded outward-facing occluded state, and an unloaded, fully open state. FIG | |
| 132 146 outward-facing protein_state These states and their connectivity can also be deduced from the calculated free-energy landscapes, which also reveal a Ca2+-loaded outward-facing occluded state, and an unloaded, fully open state. FIG | |
| 147 155 occluded protein_state These states and their connectivity can also be deduced from the calculated free-energy landscapes, which also reveal a Ca2+-loaded outward-facing occluded state, and an unloaded, fully open state. FIG | |
| 170 178 unloaded protein_state These states and their connectivity can also be deduced from the calculated free-energy landscapes, which also reveal a Ca2+-loaded outward-facing occluded state, and an unloaded, fully open state. FIG | |
| 180 190 fully open protein_state These states and their connectivity can also be deduced from the calculated free-energy landscapes, which also reveal a Ca2+-loaded outward-facing occluded state, and an unloaded, fully open state. FIG | |