LiteFold/DisProt huggingface dataset:
Protein: DP01142 (UniProt O92972)
Region: residues 2267	to 2292
Annotation: protein binding (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Upon addition of SH3 at concentrations higher than the equimolar concentration, chemical shift changes were observed for NS5A residues 200–228 (binding site B1) and 295–320 (binding site B2).
Here we have shown that NS5A interacts with SH3 domains via three distinct binding regions, with one (B3) corresponding to the canonical PxxP motif and the other two (B1 and B2) not containing such a canonical SH3 recognition element.
Residues 295-320 (B2) of the nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2267-2292 residues of the polyprotein.
Protein: DP01142 (UniProt O92972)
Region: residues 2163	to 2312
Annotation: protein binding (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
In the following, we will focus on the interaction of NS5A(191–340) with the Bin1-SH3 domain that induces the largest chemical shift changes in the NS5A spectrum. A very similar pattern of chemical shift changes and line broadening was observed in NMR titration experiments (Figure 3B) as shown in Figure 2B for the longer NS5A construct, indicating that the observed perturbation is due to direct SH3 binding events, and not to conformational changes induced by the high-affinity interaction of the SH3 domain with the PxxP motif of NS5A.
Residues 191-340 of the truncated nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2163-2312 residues of the polyprotein.
Protein: DP01142 (UniProt O92972)
Region: residues 2314	to 2340
Annotation: protein binding (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Besides the interaction with SH3 domains mediated by a canonical PxxP motif located in the LCS-2 region of NS5A (binding site B3), we were able to identify two additional peptide regions in NS5A that interact with the four SH3 domains mentioned above (Figure 2B).
For a NS5A:SH3 ratio of 1:1 (yellow), chemical shift changes can mainly be detected close to the PxxP motifs.
Here we have shown that NS5A interacts with SH3 domains via three distinct binding regions, with one (B3) corresponding to the canonical PxxP motif and the other two (B1 and B2) not containing such a canonical SH3 recognition element.
Protein: DP01142 (UniProt O92972)
Region: residues 2163	to 2341
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
More surprisingly, the hetNOE values of NS5A residues in the binding regions decrease upon interaction with SH3, in agreement with a more flexible structural ensemble. The interaction of NS5A with Bin1-SH3 via low-affinity binding sites B1 and B2 thus adds another example of a “fuzzy” complex(34-36) formed between a globular protein domain and an intrinsically disordered protein (IDP).
Protein: DP01142 (UniProt O92972)
Region: residues 2163	to 2341
Annotation: protein binding (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Consequently, at an equimolar mixture of the two proteins at 100 μM concentration, this high-affinity NS5A(191–369) binding site is saturated, and only peaks from the bound state were observable in the NMR spectra.
Residues 191-369 of the nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2163-2341 residues of the polyprotein.
Protein: DP01142 (UniProt O92972)
Region: residues 2314	to 2340
Annotation: molecular adaptor activity (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Besides the interaction with SH3 domains mediated by a canonical PxxP motif located in the LCS-2 region of NS5A (binding site B3), we were able to identify two additional peptide regions in NS5A that interact with the four SH3 domains mentioned above (Figure 2B).
Protein: DP01142 (UniProt O92972)
Region: residues 2163	to 2312
Annotation: molecular adaptor activity (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
In the following, we will focus on the interaction of NS5A(191–340) with the Bin1-SH3 domain that induces the largest chemical shift changes in the NS5A spectrum. A very similar pattern of chemical shift changes and line broadening was observed in NMR titration experiments (Figure 3B) as shown in Figure 2B for the longer NS5A construct, indicating that the observed perturbation is due to direct SH3 binding events, and not to conformational changes induced by the high-affinity interaction of the SH3 domain with the PxxP motif of NS5A.
Residues 191-340 of the truncated nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2163-2312 residues of the polyprotein.
Protein: DP01142 (UniProt O92972)
Region: residues 2267	to 2292
Annotation: molecular adaptor activity (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Here we have shown that NS5A interacts with SH3 domains via three distinct binding regions, with one (B3) corresponding to the canonical PxxP motif and the other two (B1 and B2) not containing such a canonical SH3 recognition element.
Residues 295-320 (B2) of the nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2267-2292 residues of the polyprotein.
Protein: DP01142 (UniProt O92972)
Region: residues 2172	to 2200
Annotation: molecular adaptor activity (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Upon addition of SH3 at concentrations higher than the equimolar concentration, chemical shift changes were observed for NS5A residues 200–228 (binding site B1) and 295–320 (binding site B2).
Residues 200-228 (B1) of the nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2172-2200 residues of the polyprotein.
Protein: DP01142 (UniProt O92972)
Region: residues 2305	to 2341
Annotation: molecular adaptor activity (Molecular function)
Evidence: surface plasmon resonance evidence used in manual assertion
Supporting statements:
This refers to a macroscopic system with a high binding affinity of the two proteins, consistent with a Kd of 0.1 μM measured by surface plasmon resonance for a short NS5A(333–369) fragment that comprises the entire poly-proline rich region (Fig. S6).
Residues 333-369 of the nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2305-2341 residues of the polyprotein.
Protein: DP01142 (UniProt O92972)
Region: residues 2314	to 2338
Annotation: molecular adaptor activity (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Adding a small amount of Bin1–SH3 (molar ratio of 5:1) only affects the line widths and chemical shifts of amide resonances in the region 342 to 366 (Fig. 4a) comprising three canonical SH3 binding sites (PPII helix motifs): one of class I [PP1.2 (348–354)] and two of class II [PP2.1 (343–348) and PP2.2 (351–356)] (Fig. 4c).
Residues 342-366 of the nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2314-2338 residues of the polyprotein.
Protein: DP01142 (UniProt O92972)
Region: residues 2163	to 2341
Annotation: molecular adaptor activity (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Consequently, at an equimolar mixture of the two proteins at 100 μM concentration, this high-affinity NS5A(191–369) binding site is saturated, and only peaks from the bound state were observable in the NMR spectra.
Residues 191-369 of the nonstructural protein 5A (NS5A) from Hepatitis C virus genotype 1b (strain HC-J4) corresponds to 2163-2341 residues of the polyprotein.
Protein: DP01144 (UniProt P08008)
Region: residues 1	to 100
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Backbone chemical shifts of free YopE were compared with those expected for a random coil state. The deviations from random coil values of secondary chemical shifts (Δδ) in 15N, 1H, and 13C dimensions of the chaperone binding (Cb) region of free YopE were found to be close to zero, indicating that this region was in a random coil state.
Protein: DP01144 (UniProt P08008)
Region: residues 23	to 78
Annotation: disorder to order (Structural transition)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Backbone chemical shifts of free YopE were compared with those expected for a random coil state. The deviations from random coil values of secondary chemical shifts (Δδ) in 15N, 1H, and 13C dimensions of the chaperone binding (Cb) region of free YopE were found to be close to zero, indicating that this region was in a random coil state. In contrast, when SycE was bound, these same residues of YopE had values significantly different from those typical for a random coil state. These results provide evidence for SycE promoting a transition in the Cb region from an unstructured state to a structured conformation.
Cross-references: BMRB:15878
Protein: DP01144 (UniProt P08008)
Region: residues 23	to 78
Annotation: protein binding (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
 Comparison of dually assigned residues also revealed that SycE binding had no effect on the S1 region and likewise no effect on residues 81–100 connecting the Cb region to the RhoGAP domain. These results provide direct evidence for the effect of SycE binding being strictly localized to the Cb region of YopE and not extending outside the Cb region.
The Cb region spans residues 23-78 of YopE.
Cross-references: BMRB:15878
Protein: DP01144 (UniProt P08008)
Region: residues 1	to 100
Annotation: flexible N-terminal tail (Disorder function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Backbone chemical shifts of free YopE were compared with those expected for a random coil state. The deviations from random coil values of secondary chemical shifts (Δδ) in 15N, 1H, and 13C dimensions of the chaperone binding (Cb) region of free YopE were found to be close to zero, indicating that this region was in a random coil state.
Protein: DP01144 (UniProt P08008)
Region: residues 79	to 100
Annotation: flexible linker (Disorder function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Residues 79–101 connecting the Cb region to the RhoGAP domain lacked detectable secondary structure regardless of SycE binding (Fig. 2), indicating that these residues likely serve as an unstructured linker.
Cross-references: BMRB:7426
Protein: DP01146 (UniProt O34800)
Region: residues 1	to 56
Annotation: disorder (Structural state)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
By contrast the far-UV CD spectrum of SpoIISB has less pronounced features with a shallow minimum at 200 nm. This spectrum is barely distinct from the CD spectrum of a control sample containing buffer alone. We deduce from this spectrum that SpoIISB is natively disordered.
Protein: DP01146 (UniProt O34800)
Region: residues 1	to 56
Annotation: protein binding (Molecular function)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
The addition of SpoIISB leads to an increase in the molar ellipticity of the sample at 190–200 nm and a decrease in the molar ellipticity over the wavelength range 205–225 nm. This indicates that the addition of SpoIISB to His6-CSpoIISA leads to an increase in the amount of protein that has secondary structure. Because the CD spectrum of SpoIISB alone indicates that this protein is unstructured, we conclude that CSpoIISA induces structure in SpoIISB as the two proteins form a complex.
Protein: DP01146 (UniProt O34800)
Region: residues 1	to 56
Annotation: disorder to order (Structural transition)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
The addition of SpoIISB leads to an increase in the molar ellipticity of the sample at 190–200 nm and a decrease in the molar ellipticity over the wavelength range 205–225 nm. This indicates that the addition of SpoIISB to His6-CSpoIISA leads to an increase in the amount of protein that has secondary structure. Because the CD spectrum of SpoIISB alone indicates that this protein is unstructured, we conclude that CSpoIISA induces structure in SpoIISB as the two proteins form a complex. These observations are consistent with the crystal structure, which shows that SpoIISB exhibits secondary structure in the complex that is dependent on extensive packing against SpoIISA.
Protein: DP01146 (UniProt O34800)
Region: residues 1	to 56
Annotation: protein binding (Molecular function)
Evidence: surface plasmon resonance evidence used in manual assertion
Supporting statements:
Surface plasmon resonance experiments revealed that the CSpoIISA·SpoIISB complex is stable with a dissociation constant in the nanomolar range.
Protein: DP01146 (UniProt O34800)
Region: residues 1	to 56
Annotation: protein binding (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Here, the crystal structure of a cytoplasmic fragment of SpoIISA (CSpoIISA) in complex with SpoIISB has been determined by selenomethionine-multiwavelength anomalous dispersion phasing to 2.5 Å spacing, revealing a CSpoIISA2·SpoIISB2 heterotetramer. CSpoIISA has a single domain α/β structure resembling a GAF domain with an extended α-helix at its N terminus. The two CSpoIISA protomers form extensive interactions through an intermolecular four-helix bundle. Each SpoIISB chain is highly extended and lacking tertiary structure. The SpoIISB chains wrap around the CSpoIISA dimer, forming extensive interactions with both CSpoIISA protomers.
Cross-references: PDB:3O6Q
Protein: DP01146 (UniProt O34800)
Region: residues 9	to 52
Annotation: disorder to order (Structural transition)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Here, the crystal structure of a cytoplasmic fragment of SpoIISA (CSpoIISA) in complex with SpoIISB has been determined by selenomethionine-multiwavelength anomalous dispersion phasing to 2.5 Å spacing, revealing a CSpoIISA2·SpoIISB2 heterotetramer. CSpoIISA has a single domain α/β structure resembling a GAF domain with an extended α-helix at its N terminus. The two CSpoIISA protomers form extensive interactions through an intermolecular four-helix bundle. Each SpoIISB chain is highly extended and lacking tertiary structure. The SpoIISB chains wrap around the CSpoIISA dimer, forming extensive interactions with both CSpoIISA protomers.
Cross-references: PDB:3O6Q
Protein: DP01146 (UniProt O34800)
Region: residues 1	to 56
Annotation: toxin sequestering activity (Molecular function)
Evidence: surface plasmon resonance evidence used in manual assertion
Supporting statements:
Surface plasmon resonance experiments revealed that the CSpoIISA·SpoIISB complex is stable with a dissociation constant in the nanomolar range.
High affinity binding of CSpoIISA to SpoIISB is largely conferred by the low rate of complex dissociation, reflected in the slow release of bound SpoIISB from the sensor chip after the end of the injection (Fig. 2B). This could have biological implications in effective blocking of SpoIISA by SpoIISB in vivo under conditions of limited renewal of the available SpoIISB pool in the cell.
Protein: DP01146 (UniProt O34800)
Region: residues 1	to 56
Annotation: toxin sequestering activity (Molecular function)
Evidence: x-ray crystallography evidence used in manual assertion
Supporting statements:
Here, the crystal structure of a cytoplasmic fragment of SpoIISA (CSpoIISA) in complex with SpoIISB has been determined by selenomethionine-multiwavelength anomalous dispersion phasing to 2.5 Å spacing, revealing a CSpoIISA2·SpoIISB2 heterotetramer. CSpoIISA has a single domain α/β structure resembling a GAF domain with an extended α-helix at its N terminus. The two CSpoIISA protomers form extensive interactions through an intermolecular four-helix bundle. Each SpoIISB chain is highly extended and lacking tertiary structure. The SpoIISB chains wrap around the CSpoIISA dimer, forming extensive interactions with both CSpoIISA protomers.
Protein: DP01146 (UniProt O34800)
Region: residues 1	to 56
Annotation: toxin sequestering activity (Molecular function)
Evidence: experimental phenotypic evidence used in manual assertion
Supporting statements:
Inappropriate spoIISA expression causes lysis of vegetatively growing B. subtilis cells and Escherichia coli cells when expressed heterologously, effects that are countered by co-expression of spoIISB, identifying SpoIISA-SpoIISB as a toxin-antitoxin system.
Protein: DP01146 (UniProt O34800)
Region: residues 13	to 53
Annotation: toxin sequestering activity (Molecular function)
Evidence: deletion mutation phenotypic evidence used in manual assertion
Supporting statements:
However, when spoIISA was co-expressed with spoIISB deletants encoding proteins in which 3 or 12 N-terminal residues were replaced by Met-Ala and Met-Ala-Ala, respectively, the sporulation efficiencies of the strains were similar to the wild type strain (2.9 × 108 and 8.4 × 108 spores/ml; Fig. 4A). These results suggest that the N-terminal 12 residues are dispensable for the antitoxin activity of SpoIISB in vivo. Because the missing residues include Cys9′, the disulfide bridge is probably not critical for the antidote function. The sporulation efficiencies of strains encoding SpoIISB proteins with C-terminal deletions of 16, 12, 8, and 4 residues were 9.5 × 103, 5.7 × 103, 6.6 × 103, and 4.8 × 103 spores/ml, respectively. These are similar to the sporulation efficiency of cells in which the entire spoIISB gene has been deleted. Thus, C-terminal deletion of four or more residues leads to a loss of the capacity of SpoIISB to overcome SpoIISA toxicity.
Protein: DP01147 (UniProt Q9I322)
Region: residues 1	to 81
Annotation: disorder (Structural state)
Evidence: cleavage assay evidence used in manual assertion
Supporting statements:
Whereas ExsE is rapidly degraded to small undetectable peptides by SDS-PAGE following a brief treatment with subtilisin. The limited stability of ExsE towards proteolytic digestion suggests that it either lacks of stable tertiary fold or adopts a highly dynamic structure.
Protein: DP01147 (UniProt Q9I322)
Region: residues 1	to 81
Annotation: disorder (Structural state)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
ExsE lacks stable secondary structures, and resembles random coil conformations in solution based on circular dichroism.
Protein: DP01147 (UniProt Q9I322)
Region: residues 1	to 81
Annotation: molecular function regulator (Molecular function)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
ExsE interacts with the secretion chaperone ExsC.
Protein: DP01147 (UniProt Q9I322)
Region: residues 1	to 81
Annotation: protein binding (Molecular function)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
ExsE interacts with the secretion chaperone ExsC.
Protein: DP01147 (UniProt Q9I322)
Region: residues 1	to 81
Annotation: disorder to order (Structural transition)
Evidence: far-UV circular dichroism evidence used in manual assertion
Supporting statements:
We identified regions that are partially ordered in the largely unstructured ExsE polypeptide. We found that these regions are correlated with those that adopt secondary structures when ExsE binds to ExsC. In the stable ExsE-ExsC interface, ExsE is in the fully extended conformation, and two signature β strands at both the N- and C-termini of the ExsE polypeptide pair with two symmetry-related five-stranded β sheets of dimeric ExsC. These findings suggest the importance of pre-existing ordered structured in binding of intrinsically disordered proteins to their targets.
Protein: DP01147 (UniProt Q9I322)
Region: residues 1	to 81
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
The 1H-15N HSQC NMR spectra showed that the amide proton chemical shifts of ExsE collapse in a narrow range of 7.8-8.6 ppm, consistent with the absence of secondary structures in ExsE. ExsE is highly dynamic throughout the polypeptide chain, as indicated by backbone dynamic analysis on relaxation times T1 and T2 of 15N and the {1H}-15N heteronuclear NOEs. Some local regions in the globally unstructured ExsE may possess residual or transient structure.
Protein: DP01148 (UniProt A0A068MVV3)
Region: residues 1	to 65
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
The 1H–15N HSQC spectrum showed signals with narrow lines and limited dispersion. In total, the δ2D prediction suggests that 29% of the residues in IF7 adopt helical conformations (helical percentages of >10%) and 71% of them populate random-coil conformations.
Cross-references: BMRB:25921
Protein: DP01148 (UniProt A0A068MVV3)
Region: residues 1	to 65
Annotation: protein binding (Molecular function)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
Glutamine synthetase (GS) is controlled by a post-translational regulatory mechanism involving protein–protein interactions with 65-residue (IF7) and 149-residue (IF17) inactivating proteins.
Protein: DP01148 (UniProt A0A068MVV3)
Region: residues 1	to 65
Annotation: disorder (Structural state)
Evidence: nuclear magnetic resonance spectroscopy evidence used in manual assertion
Supporting statements:
The NOE values of residues of the isolated IF7 were below 0.52 (the value observed for Ala29) (Fig. 1 D). Keeping in mind that at 11.7 T the maximum value for the NOE is 0.70 [52] in an isotropically tumbling molecule without internal motions, we must conclude that isolated IF7 had a highly flexible backbone.
Protein: DP01148 (UniProt A0A068MVV3)
Region: residues 1	to 65
Annotation: disorder (Structural state)
Evidence: small-angle X-ray scattering evidence used in manual assertion
Supporting statements:
We did not observe a well-defined peak in the Kratky plot of isolated IF7 data, proving that the protein did not have a globular shape. In a first approximation, SAXS data for IF7 were fitted by using either the Guinier or Debye equation (Fig. S2). The two approaches provided a radius of gyration Rg = 24.2 ± 0.2 Å (in the Guinier approach) and Rg = 26.95 ± 0.05 Å (in the Debye approach). The Debye equation is more appropriate for disordered chains than the Guinier one, and accurately fitted our experimental data, as shown by the continuous line (Fig. 3).