bluuebunny/crossref_metadata_2025_split huggingface dataset:
Are proteins ideal mixtures of amino acids? Analysis of energy parameter sets
Authors: Godzik Adam , Skolnick Jeffrey , Koliński Andrzej 
Published: February 2009

AbstractVarious existing derivations of the effective potentials of mean force for the two‐body interactions between amino acid side chains in proteins are reviewed and compared to each other. The differences between different parameter sets can be traced to the reference state used to define the zero of energy. Depending on the reference state, the transfer free energy or other pseudo‐one‐body contributions can be present to various extents in two‐body parameter sets. It is, however, possible to compare various derivations directly by concentrating on the “excess” energy— a term that describes the difference between a real protein and an ideal solution of amino acids. Furthermore, the number of protein structures available for analysis allows one to check the consistency of the derivation and the errors by comparing parameters derived from various subsets of the whole database. It is shown that pair interaction preferences are very consistent throughout the database. Independently derived parameter sets have correlation coefficients on the order of 0.8, with the mean difference between equivalent entries of 0.1kT. Also, the low‐quality (low resolution, little or no refinement) structures show similar regularities. There are, however, large differences between interaction parameters derived on the basis of crystallographic structures and structures obtained by the NMR refinement. The origin of the latter difference is not yet understood.
Cα‐based torsion angles: A simple tool to analyze protein conformational changes
Authors: Flocco Maria M. , Mowbray Sherry L. 
Published: February 2009

AbstractA simple method is presented for the analysis of protein conformational changes based on the comparison of torsion angles defined by four consecutive Cα atoms. The technique was applied successfully to proteins that undergo hinge motion and shear motion. In the case of both MBP and LAO, which represent examples of hinge motion, the plot of the differences in Cα‐torsion angles between the open and closed forms of the proteins helped us to formulate a more thorough description of the conformational change: a large displacement of one domain with respect to the other where one of the domains does not behave like a rigid body but exhibits some degree of flexibility. The analysis of citrate synthase, which is an example of shear motion, shows that the largest differences in Cα‐torsion angles between the open and closed conformations are clustered around residues that belong to segments connecting α‐helices, whereas the helices themselves appear to be rigid; this is in agreement with previous results obtained by detailed least‐squares superpositions (Lesk AM, Chothia C, 1984, J Mol Biol 174:175–191).
Relative effectiveness of various anions on the solubility of acidicHypoderma lineatumcollagenase at pH 7.2
Authors: Carbonnaux Claire , Ries‐Kautt Madeleine , Ducruix Arnaud 
Published: February 2009

AbstractThe effects of various anions on decreasing the solubility of acidicHypoderma lineatumcollagenase at pH 7.2 and 18 °C were qualitatively defined by replacing the crystallizing agent of known crystallization conditions by various ammonium salts. The solubility curves measured in the presence of the sulfate, phosphate, citrate, and chloride ammonium salts gave the following ranking of anions: HPO42‐/H2PO4−> SO42‐> citrate3‐/citrate2‐>> Cl−. This order is in agreement with the Hofmeister series. In a previous study on the solubility at pH 4.5 of lysozyme, a basic protein, the effectiveness of anions in decreasing the solubility was found to be in the reverse order. This suggests that the effectiveness of anions in the crystallization of proteins is dependent on the net charge of the protein, i.e., depending on whether a basic protein is crystallized at acidic pH or an acidic protein at basic pH.
Denaturant m values and heat capacity changes: Relation to changes in accessible surface areas of protein unfolding
Authors: Myers Jeffrey K. , Nick Pace C. , Martin Scholtz J. 
Published: February 2009

AbstractDenaturant m values, the dependence of the free energy of unfolding on denaturant concentration, have been collected for a large set of proteins. The m value correlates very strongly with the amount of protein surface exposed to solvent upon unfolding, with linear correlation coefficients of R = 0.84 for urea and R = 0.87 for guanidine hydrochloride. These correlations improve to R = 0.90 when the effect of disulfide bonds on the accessible area of the unfolded protein is included. A similar dependence on accessible surface area has been found previously for the heat capacity change (ΔCp), which is confirmed here for our set of proteins. Denaturant m values and heat capacity changes also correlate well with each other. For proteins that undergo a simple two‐state unfolding mechanism, the amount of surface exposed to solvent upon unfolding is a main structural determinant for both m values and ΔCp.
The role of glutamate 87 in the kinetic mechanism of Thermus thermophilus isopropylmalate dehydrogenase
Authors: Dean Antony M. , Dvorak Laura 
Published: February 2009

AbstractThe kinetic mechanism of the oxidative decarboxylation of 2R,3S‐isopropylmalate by the NAD‐dependent isopropylmalate dehydrogenase of Thermus thermophilus was investigated. Initial rate results typical of random or steady‐state ordered sequential mechanisms are obtained for both the wild‐type and two mutant enzymes (E87G and E87Q) regardless of whether natural or alternative substrates (2R‐malate, 2R,3S‐tartrate and/or NADP) are utilized. Initial rate data fail to converge on a rapid equilibrium‐ordered pattern despite marked reductions in specificity (kcat/Km) caused by the mutations and alternative substrates. Although the inhibition studies alone might suggest an ordered kinetic mechanism with cofactor binding first, a detailed analysis reveals that the expected noncompetitive patterns appear uncompetitive because the dissociation constants from the ternary complexes are far smaller than those from the binary complexes. Equilibrium fluorescence studies both confirm the random binding of substrates and the kinetic estimates of the dissociation constants of the substrates from the binary complexes. The latter are not disturbed markedly by the mutations at site 87. Mutations at site 87 do not affect the dissociation constants from the binary complexes, but do greatly increase the Michaelis constants, indicating that E87 helps stabilize the Michaelis complex of the wild‐type enzyme. The available structural data, the patterns of the kinetics results, and the structure of a pseudo‐Michaelis complex of the homologous isocitrate dehydrogenase of Escherichia coli suggest that E87 interacts with the nicotinamide ring.
Molecular modeling studies of HIV‐1 reverse transcriptase nonnucleoside inhibitors: Total energy of complexation as a predictor of drug placement and activity
Authors: Kroeger Smith Marilyn B. , Hughes Stephen H. , Boyer Paul L. , Michejda Christopher J. , Rouzer Carol A. , Taneyhill Lisa A. , Smith Richard H. , Smith Nathan A. , Janssen Paul A.J. , Moereels Henri , Koymans Luc , Arnold Edward , Ding Jianping , Das Kalyan , Zhang Wanyi 
Published: February 2009

AbstractComputer modeling studies have been carried out on three nonnucleoside inhibitors complexed with human immunodeficiency virus type 1 (HIV‐1) reverse transcriptase (RT), using crystal coordinate data from a subset of the protein surrounding the binding pocket region. Results from the minimizations of solvated complexes of 2‐cyclopropyl‐4‐methyl‐5,11‐dihydro‐5H‐dipyrido[3,2‐b:2′,3′‐e][1,4]diazepin‐6‐one (nevirapine), α‐anilino‐2, 6‐dibromophenylacetamide (α‐APA), and 8‐chloro‐tetrahydro‐imidazo(4,5,1‐jk)(1,4)‐benzodiazepin‐2(1H)‐thione (TIBO) show that all three inhibitors maintain a very similar conformational shape, roughly overlay each other in the binding pocket, and appear to function as π‐electron donors to aromatic side‐chain residues surrounding the pocket. However, side‐chain residues adapt to each bound inhibitor in a highly specific manner, closing down around the surface of the drug to make tight van der Waals contacts. Consequently, the results from the calculated minimizations reveal that only when the inhibitors are modeled in a site constructed from coordinate data obtained from their particular RT complex can the calculated binding energies be relied upon to predict the correct orientation of the drug in the pocket. In the correct site, these binding energies correlate with EC50values determined for all three inhibitors in our laboratory. Analysis of the components of the binding energy reveals that, for all three inhibitors, solvation of the drug is endothermic, but solvation of the protein is exothermic, and the sum favors complex formation. In general, the protein is energetically more stable and the drug less stable in their complexes as compared to the reactant conformations. For all three inhibitors, interaction with the protein in the complex is highly favorable. Interactions of the inhibitors with individual residues correlate with crystallographic and site‐specific mutational data. π‐Stacking interactions are important in binding and correlate with drug HOMO RHF/6–31G* energies. Modeling results are discussed with respect to the mechanism of complex formation and the design of nonnucleoside inhibitors that will be more effective against mutants of HIV‐1 RT that are resistant to the currently available drugs.
The Alacoil: A very tight, antiparallel coiled‐coil of helices
Authors: Gernert Kim M. , Surles Mark C. , Labean Thomas H. , Richardson Jane S. , Richardson David C. 
Published: February 2009

AbstractThe Alacoil is an antiparallel (rather than the usual parallel) coiled‐coil of α‐helices with Ala or another small residue in every seventh position, allowing a very close spacing of the helices (7.5–8.5 Å between local helix axes), often over four or five helical turns. It occurs in two distinct types that differ by which position of the heptad repeat is occupied by Ala and by whether the closest points on the backbone of the two helices are aligned or are offset by half a turn. The aligned, or ROP, type has Ala in position “d” of the heptad repeat, which occupies the “tip‐to‐tip” side of the helix contact where the Cα–Cβ bonds point toward each other. The more common offset, or ferritin, type of Alacoil has Ala in position “a” of the heptad repeat (where the Cα‐Cβ bonds lie back‐to‐back, on the “knuckle‐touch” side of the helix contact), and the backbones of the two helices are offset vertically by half a turn. In both forms, successive layers of contact have the Ala first on one and then on the other helix.The Alacoil structure has much in common with the coiled‐coils of fibrous proteins or leucine zippers: both are α‐helical coiled‐coils, with a critical amino acid repeated every seven residues (the Leu or the Ala) and a secondary contact position in between. However, Leu zippers are between aligned, parallel helices (often identical, in dimers), whereas Alacoils are between antiparallel helices, usually offset, and much closer together. The Alacoil, then, could be considered as an “Ala anti‐zipper.” Leu zippers have a classic “knobs‐into‐holes” packing of the Leu side chain into a diamond of four residues on the opposite helix; for Alacoils, the helices are so close together that the Ala methyl group must choose one side of the diamond and pack inside a triangle of residues on the other helix.We have used the ferritin‐type Alacoil as the basis for the de novo design of a 66‐residue, coiled helix hairpin called “Alacoilin.” Its sequence is: cmSPDQWDKE AAQYDAHAQE FEKKSHRNng TPEADQYRHM ASQY QAMAQK LKAIANQLKK Gseter (with “a” heptad positions underlined and nonhelical parts in lowercase), which we will produce and test for both stability and uniqueness of structure.
Covariation of residues in the homeodomain sequence family
Authors: Clarke Neil D. 
Published: February 2009

AbstractHomeodomains are 60 amino acid DNA binding domains found in numerous eukaryotic transcription factors. The homeodomain family is a useful system for studying sequence‐structure relationships because several hundred sequences are known and the structures of several homeodomains have been determined. Covariation of amino acid residues in the homeodomain family has been investigated to see whether strongly covariant residue pairs can be understood in terms of the structure and function of these domains. Among 16 strongly covariant pairs examined, 2 are explained by the ability to form salt bridges, and 9 appear related to the DNA binding function of the proteins. For the remaining 5 pairs, the rationale for covariance remains unclear and the likelihood of artifactual correlations is discussed in the context of experimental and evolutionary biases in the selection of sequences. No significant correlation was found between covariance and structural proximity in the hydrophobic core.
Drosophilaengrailed‐1, 10‐phenanthroline chimeras as probes of homeodomain‐DNA complexes
Authors: Pan Clark Q. , Landgraf Ralf , Sigman David S. 
Published: February 2009

AbstractWe have converted theDrosophilaengrailed homeodomain into a sequence‐specific nuclease by linking the protein to the chemical nuclease 1,10‐phenanthroline‐copper (OP‐Cu). Unique cysteines were introduced at six positions into the homeodomain by site‐directed mutagenesis for the covalent attachment of OP‐Cu. The varied DNA‐binding affinity and specificity of these mutants and the DNA cleavage pattern of their OP‐Cu derivatives allowed us to assess the crystal structure of the engrailed homeodomain‐DNA complex. We have also achieved site‐specific double‐stranded DNA scission with one of the homeodomain mutants, E28C, which has the potential of being used to identify engrailed binding sites in the genome. Because the homeodomain is so well conserved among members of the homeodomain‐containing protein family, other homeodomain proteins can be converted into nucleases by attaching OP‐Cu at position 28 of their homeodomains.
Comparison of the accuracy of protein solution structures derived from conventional and network‐edited NOESY data
Authors: Hoogstraten Charles G. , Choe Soheui , Westler William M. , Markley John L. 
Published: February 2009

AbstractNetwork‐editing experiments are variants of the basic NOESY experiment that allow more accurate direct measurement of interproton distances in macromolecules by defeating specific spin‐diffusion pathways. Two network‐editing approaches, block‐decoupled NOESY and complementary‐block‐decoupled‐NOESY, were applied as three‐dimensional, heteronuclear‐edited experiments to distance measurement in a small protein, turkey ovomucoid third domain (OMTKY3). Two‐hundred and twelve of the original 655 distance constraints observed in this molecule (Krezel AM et al., 1994, J Mol Biol 242:203–214) were improved by their replacement by distances derived from network‐edited spectra, and distance geometry/simulated annealing solution structure calculations were performed from both the unimproved and improved distance sets. The resulting two families of structures were found to differ significantly, the most important differences being the hinge angle of a β‐turn and an expansion of the sampled conformation space in the region of the reactive‐site loop. The structures calculated from network‐editing data are interpreted as a more accurate model of the solution conformation of OMTKY3.
The use of side‐chain packing methods in modeling bacteriophage repressor and cro proteins
Authors: Chung Su Yun , Subbiah S. 
Published: February 2009

AbstractIn recent years, it has been repeatedly demonstrated that the coordinates of the main‐chain atoms alone are sufficient to determine the side‐chain conformations of buried residues of compact proteins. Given a perfect backbone, the side‐chain packing method can predict the side‐chain conformations to an accuracy as high as 1.2 Å RMS deviation (RMSD) with greater than 80% of the χ angles correct. However, similarly rigorous studies have not been conducted to determine how well these apply, if at all, to the more important problem of homology modeling per se. Specifically, if the available backbone is imperfect, as expected for practical application of homology modeling, can packing constraints alone achieve sufficiently accurate predictions to be useful? Here, by systematically applying such methods to the pairwise modeling of two repressor and two cro proteins from the closely related bacteriophages 434 and P22, we find that when the backbone RMSD is 0.8 Å, the prediction on buried side chain is accurate with an RMS error of 1.8 Å and approximately 70% of the χ angles correctly predicted. When the backbone RMSD is larger, in the range of 1.6–1.8 Å, the prediction quality is still significantly better than random, with RMS error at 2.2 Å on the buried side chains and 60% accuracy on χ angles. Together these results suggest the following rules‐of‐thumb for homology modeling of buried side chains. When the sequence identity between the modeled sequence and the template sequence is >50% (or, equivalently, the expected backbone RMSD is <1 Å), side‐chain packing methods work well. When sequence identity is between 30–50%, reflecting a backbone RMS error of 1–2 Å, it is still valid to use side‐chain packing methods to predict the buried residues, albeit with care. When sequence identity is below 30% (or backbone RMS error greater than 2 Å), the backbone constraint alone is unlikely to produce useful models. Other methods, such as those involving the use of database fragments to reconstruct a template backbone, may be necessary as a complementary guide for modeling.
Fluorescence of native single‐Trp mutants in the lactose permease from Escherichia coli: Structural properties and evidence for a substrate‐induced conformational change
Authors: Weitzman Cindy , Consler Thomas G. , Kaback H. Ronald 
Published: February 2009

AbstractSix single‐Trp mutants were engineered by individually reintroducing each of the native Trp residues into a functional lactose permease mutant devoid of Trp (Trp‐less permease; Menezes ME, Roepe PD, Kaback HR, 1990, Proc Natl Acad Sci USA 87:1638–1642), and fluorescent properties were studied with respect to solvent accessibility, as well as alterations produced by ligand binding. The emission of Trp 33, Trp 78, Trp 171, and Trp 233 is strongly quenched by both acrylamide and iodide, whereas Trp 151 and Trp 10 display a decrease in fluorescence in the presence of acrylamide only and no quenching by iodide. Of the six single‐Trp mutants, only Trp 33 exhibits a significant change in fluorescence (ca. 30% enhancement) in the presence of the substrate analog β,D‐galactopyranosyl 1‐thio‐β,D‐galactopyranoside (TDG). This effect was further characterized by site‐directed fluorescent studies with purified single‐Cys W33 → C permease labeled with 2‐(4′‐maleimidylanilino)‐naphthalene‐6‐sulfonic acid (MIANS). Titration of the change in the fluorescence spectrum reveals a 30% enhancement accompanied with a 5‐nm blue shift in the emission maximum, and single exponential behavior with an apparent KD of 71 μM. The effect of substrate binding on the rate of MIANS labeling of single‐Cys 33 permease was measured in addition to iodide and acrylamide quenching of the MIANS‐labeled protein. Complete blockade of labeling is observed in the presence of TDG, as well as a 30% decrease in accessibility to iodide with no change in acrylamide quenching. Overall, the findings are consistent with the proposal (Wu J, Frillingos S, Kaback HR, 1995a, Biochemistry 34:8257–8263) that ligand binding induces a conformational change at the C‐terminus of helix I such that Pro 28 and Pro 31, which are on one face, become more accessible to solvent, whereas Trp 33, which is on the opposite face, becomes less accessible to the aqueous phase. The findings regarding accessibility to collisional quenchers are also consistent with the predicted topology of the six native Trp residues in the permease.
Two steps in the transition between the native and acid states of bovine α‐lactalbumin detected by circular polarization of luminescence: Evidence for a premolten globule state?
Authors: Gussakovsky Eugene E. , Haas Elisha 
Published: February 2009

AbstractA few studies indirectly support the existence of an intermediate in the transition of Ca2+‐saturated bovine α‐lactalbumin (α‐LA) from the native (N) to the acidic (A) state, known as the molten globule state. However, direct experimental evidence for the appearance of this intermediate has not been obtained. The signal of circular polarization of luminescence (CPL) is sensitive to fine conformational transitions because of its susceptibility to changes in the environmental asymmetry of fluorescent chromophores in their excited electronic states. In the present study, CPL measurements were applied using the intrinsic tryptophan fluorescence of α‐LA as well as the fluorescence of 8‐anilino‐1‐naphthalenesulfonic acid (ANS) bound to α‐LA. CPL of tryptophan and ANS was measured in the pH range of 2.5–6 in order to find direct experimental evidence for the proposed intermediate. CPL (characterized by the emission anisotropy factor, gem) depends on the asymmetry of the protein molecular structure in the environment of the tryptophan and the ANS chromophores in the excited electronic state. The pH dependence of both the gab, absorption anisotropy factor determined by CD, and the ANS steady state fluorescence, showed a single transition at pH 3–3.7 as already reported elsewhere. This transition was interpreted as being a result of a change of the α‐LA tertiary structure, which resulted in a loss of asymmetry of the environment of both the tryptophan residues and the ANS hydrophobic binding sites. The pH dependence of the tryptophan and ANS gem showed an additional conformational transition at pH 4–5, which coincided with the pKa of Ca2+ dissociation (pKa 5), as predicted by Permyakov et al. (1981, Biochem Biophys Res Commun 100:191–197). The titration curve showed that there is a pH range between 3.7 and 4.1 in which α‐LA exists in an intermediate state between the N‐ and A‐state. We suggest that the intermediate is the premolten globule state characterized by a reduced Ca2+ binding to the α‐LA, native‐like tertiary structure, and reduced asymmetric fluctuation of the tertiary structure on the nanosecond time scale. This intermediate resembles the “critical activated state” theoretically deduced by Kuwajima et al. (1989, J Mol Biol 206:547–561). The present study demonstrates the power of CPL measurements for the investigation of folding/unfolding transitions in proteins.
Crystal structure of the dipeptide binding protein from Escherichia coli involved in active transport and chemotaxis
Authors: Dunten Pete , Mowbray Sherry L. 
Published: February 2009

AbstractThe Escherichia coli periplasmic dipeptide binding protein functions in both peptide transport and taxis toward peptides. The structure of the dipeptide binding protein in complex with Gly‐Leu (glycyl‐L‐leucine) has been determined at 3.2 Å resolution. The binding site for dipeptides is designed to recognize the ligand's backbone while providing space to accommodate a variety of side chains. Some repositioning of protein side chains lining the binding site must occur when the dipeptide's second residue is larger than leucine. The protein's fold is very similar to that of the Salmonella typhimurium oligopeptide binding protein, and a comparison of the structures reveals the structural basis for the dipeptide binding protein's preference for shorter peptides.