Inherent chirality of the retinal chromophore in rhodopsin-A nonempirical theoretical analysis of chiroptical data

CASSCF and GAUSSIAN CIS calculations were performed on ground and excited states of different conformations of 11-cis-retinal protonated Schiff bases, the chromophore of rhodopsin, in order to study their chiroptical properties and attempt a correlation between absolute conformation and CD-spectral data. Geometries were taken from molecular models, from published rhodopsin models, and from retinal conformations obtained from molecular dynamics with geometry restraints. In all the cases studied we find that a positive sense of twist about the C12-C13 bond correlates with a calculated positive CD of the long wavelength absorption band; the twist of the C6-C7 bond modulates this primary contribution of the C12-C13 bond. The correlation of the beta-band with structural features is not straightforward. Calculations on bathorhodopsin lend support to the idea that this intermediate is a highly twisted all-trans-conformation.     

A genetic algorithm that seeks native states of peptides and proteins.

We describe a computer algorithm to predict native structures of proteins and peptides from their primary sequences, their known native radii of gyration, and their known disulfide bonding patterns, starting from random conformations. Proteins are represented as simplified real-space main chains with single-bead side chains. Nonlocal interactions are taken from structural database-derived statistical potentials, as in an earlier treatment. Local interactions are taken from simulations of (phi, psi) energy surfaces for each amino acid generated using the Biosym Discover program. Conformational searching is done by a genetic algorithm-based method. Reasonable structures are obtained for melittin (a 26-mer), avian pancreatic polypeptide inhibitor (a 36-mer), crambin (a 46-mer), apamin (an 18-mer), tachyplesin (a 17-mer), C-peptide of ribonuclease A (a 13-mer), and four different designed helical peptides. A hydrogen bond interaction was tested and found to be generally unnecessary for helical peptides, but it helps fold some sheet regions in these structures. For the few longer chains we tested, the method appears not to converge. In those cases, it appears to recover native-like secondary structures, but gets incorrect tertiary folds.     

Complete Amino Acid Sequence of Chitinase-a from Bulbs of Gladiolus (Gladiolus gandavensis)

The complete amino acid sequence of gladiolus bulb chitinase-a (GBC-a) was determined. First the tryptic peptides from GBC-a after it was reduced and S-carboxymethylated were sequenced and then the peptides were further studied by chemical cleavage of the enzyme. GBC-a consisted of 274 amino acid residues and had a molecular mass of 30,714 Da. Two consensus sequences essential for chitinase activity by plant class III chitinases were conserved in GBC-a, although its sequence similarity with plant class III chitinases was less than 20%. Sequence comparison of GBC-a with sequences of other proteins in a protein identification resource (PIR) showed that the GBC-a sequence was 33% similar to that of narbonin, a seed storage 2S globulin from narbon beans.     

Structural compromise of disallowed conformations in peptide and protein structures

Using a data set of 454 crystal structures of peptides and 80 crystal structures of non-homologous proteins solved at ultra high resolution of 1.2 A or better we have analyzed the occurrence of disallowed Ramachandran (phi, psi) angles. Out of 1492 and 13508 non-glycyl residues in peptides and proteins respectively 12 and 76 residues in the two datasets adopt clearly disallowed combinations of Ramachandran angles. These examples include a number of conformational points which are far away from any of the allowed regions in the Ramachandran map. According to the Ramachandran map a given (phi, psi) combination is considered disallowed when two non-bonded atoms in a system of two-linked peptide units with ideal geometry are prohibitively proximal in space. However, analysis of the disallowed conformations in peptide and protein structures reveals that none of the observations of disallowed conformations in the crystal structures correspond to a short contact between non-bonded atoms. A further analysis of deviations of bond lengths and angles, from the ideal peptide geometry, at the residue positions of disallowed conformations in the crystal structures suggest that individual bond lengths and angles are all within acceptable limits. Thus, it appears that the rare tolerance of disallowed conformations is possible by gentle and acceptable deviations in a number of bond lengths and angles, from ideal geometry, over a series of bonds resulting in a net gross effect of acceptable non-bonded inter-atomic distances.     

Not Only Expansion: Proline Content and Density Also Induce Disordered Protein Conformation Compaction

Intrinsically disordered proteins (IDPs) adopt a wide array of different conformations that can be constrained by the presence of proline residues, which are frequently found in IDPs. To assess the effects of proline, we designed a series of peptides that differ with respect to the number of prolines in the sequence and their organization. Using high-resolution atomistic molecular dynamics simulations, we found that accounting for whether the proline residues are clustered or isolated contributed significantly to explaining deviations in the experimentally-determined gyration radii of IDPs from the values expected based on the Flory scaling-law. By contrast, total proline content makes smaller contribution to explaining the effect of prolines on IDP conformation. Proline residues exhibit opposing effects depending on their organizational pattern in the IDP sequence. Clustered prolines (i.e., prolines with ≤2 intervening non-proline residues) result in expanded peptide conformations whereas isolated prolines (i.e., prolines with >2 intervening non-proline residues) impose compacted conformations. Clustered prolines were estimated to induce an expansion of ∼20% in IDP dimension (via formation of PPII structural elements) whereas isolated prolines were estimated to induce a compaction of ∼10% in IDP dimension (via the formation of backbone turns). This dual role of prolines provides a mechanism for conformational switching that does not rely on the kinetically much slower isomerization of cis proline to the trans form. Bioinformatic analysis demonstrates high populations of both isolated and clustered prolines and implementing them in coarse-grained molecular dynamics models illustrates that they improve the characterization of the conformational ensembles of IDPs.     

Observation of residual dipolar couplings in short peptides

Residual dipolar couplings provide information on the orientation of individual bond vectors with respect to a unique set of molecular axes. We report that short peptides from 2 to 15 amino acids in length of arbitrary sequence exhibit a modest range of residual dipolar couplings when aligned in either strained polyacrylamide gels or alkyl-PEG bicelles. The absence of significant line broadening in gels suggests peptides align predominantly through steric interactions with the polyacrylamide matrix. However, broadening of NMR lines for a subset of residues aligned in bicelles indicates some peptides bind weakly to these lipid disks, yet a weak negative correlation between the couplings measured in gels and bicelles is consistent with steric hindrance playing a role in both media. The observation of dipolar couplings for peptides of length 10-15 suggests the statistical segment lengths of polypeptide chains must often be >10-15 residues, with data from denatured proteins indicating even larger values. Presumably, local side-chain backbone interactions severely restrict chain flexibility, with the cumulative effect of many such restrictions giving rise to biases in chain direction that may persist for the entire length of a protein chain. Comparison of experimental dipolar couplings for peptides with couplings calculated for ensembles of conformations generated by molecular dynamics should permit evaluation of the accuracy of molecular mechanics potentials in reproducing sequence-specific preferences for phi and psi angles.     

Comparative structural analyses of corticosteroid binding globulin

Some physicochemical characteristics of corticosteroid binding globulin (CBG) in several species have been determined. Molecular radii were determined from Ferguson plots and were used in conjunction with sedimentation coefficients determined by sucrose density gradient centrifugation to calculate the molecular weights of the CBG. These were found to range from 44,200 (dog) to 60,000 (turtle) for most species. The squirrel monkey was found to have a molecular weight twice that of other species (119,800). Purified CBG was prepared from human, rat, and guinea pig sera. The molecular weights of the purified material, as determined by gel electrophoresis in the presence of sodium dodecyl sulfate, were in excellent agreement with those determined by Ferguson analysis. Careful examination of the purified proteins by electrophoresis at pH 8.3 revealed that each consisted of two closely related electrophoretic variants. Tryptic peptides were prepared from the purified proteins and separated by reversed phase HPLC chromatography. The peptide patterns were identical for the three proteins with the exception of three hydrophilic peptides. Amino terminal sequence analysis of the rat and human proteins revealed no apparent homology, however. The immunologic relatedness of the three purified proteins was also examined, but no crossreactivity was observed. The results obtained suggest that while the molecular size and hydrophobicity of peptides have been conserved across species considerable surface differences must exist.     

Recognition in cell adhesion. A comparative study of the conformations of RGD-containing peptides by Monte Carlo and NMR methods

Many of the proteins that mediate cell adhesion processes processes-fibronectin, fibrinogen, vitronectin, von Willebrand factor, osteopontin, laminin and various collagens--contain the amino acid sequence Arg-Gly-Asp. Short peptides that include this sequence have been shown to inhibit the interactions of cell adhesion proteins with their receptors and to have dramatic effects on developmental processes involving cellular recognition. In order to determine which conformations are accessible to Arg-Gly-Asp containing peptides, we analyzed tri-, tetra- and pentapeptides using molecular mechanics and Monte Carlo methods, and studied the solution conformations of the pentapeptide Gly-Arg-Gly-Asp-Ser using nuclear magnetic resonance techniques. The Monte Carlo method was used to: (a) identify the low energy conformations of the peptides and (b) evaluate their thermodynamic properties. In the case of the pentapeptide Gly-Arg-Gly-Asp-Gly, the four stable conformations include three with reverse turns and one open structure. The conformations found in this analysis are compatible with the nuclear magnetic resonance (nuclear Overhauser effect) data.     

Do all backbone polar groups in proteins form hydrogen bonds?

Evidence from proteins and peptides supports the conclusion that intrapeptide hydrogen bonds stabilize the folded form of proteins. Paradoxically, evidence from small molecules supports the opposite conclusion, that intrapeptide hydrogen bonds are less favorable than peptide-water hydrogen bonds. A related issue-often lost in this debate about comparing peptide-peptide to peptide- water hydrogen bonds-involves the energetic cost of an unsatisfied hydrogen bond. Here, experiment and theory agree that breaking a hydrogen bond costs between 5 and 6 kcal/mol. Accordingly, the likelihood of finding an unsatisfied hydrogen bond in a protein is insignificant. This realization establishes a powerful rule for evaluating protein conformations.     

Modeling protein loops using a phi i + 1, psi i dimer database.

We present an automated method for modeling backbones of protein loops. The method samples a database of phi i + 1 and psi i angles constructed from a nonredundant version of the Protein Data Bank (PDB). The dihedral angles phi i + 1 and psi i completely define the backbone conformation of a dimer when standard bond lengths, bond angles, and a trans planar peptide configuration are used. For the 400 possible dimers resulting from 20 natural amino acids, a list of allowed phi i + 1, psi i pairs for each dimer is created by pooling all such pairs from the loop segments of each protein in the nonredundant version of the PDB. Starting from the N-terminus of the loop sequence, conformations are generated by assigning randomly selected pairs of phi i + 1, psi i for each dimer from the respective pool using standard bond lengths, bond angles, and a trans peptide configuration. We use this database to simulate protein loops of lengths varying from 5 to 11 amino acids in five proteins of known three-dimensional structures. Typically, 10,000-50,000 models are simulated for each protein loop and are evaluated for stereochemical consistency. Depending on the length and sequence of a given loop, 50-80% of the models generated have no stereochemical strain in the backbone atoms. We demonstrate that, when simulated loops are extended to include flanking residues from homologous segments, only very few loops from an ensemble of sterically allowed conformations orient the flanking segments consistent with the protein topology. The presence of near-native backbone conformations for loops from five different proteins suggests the completeness of the dimeric database for use in modeling loops of homologous proteins. Here, we take advantage of this observation to design a method that filters near-native loop conformations from an ensemble of sterically allowed conformations. We demonstrate that our method eliminates the need for a loop-closure algorithm and hence allows for the use of topological constraints of the homologous proteins or disulfide constraints to filter near-native loop conformations.     

Empirical solvation models can be used to differentiate native from near-native conformations of bovine pancreatic trypsin inhibitor.

Several hydration models for peptides and proteins based on solvent accessible surface area have been proposed previously. We have evaluated some of these models as well as four new ones in the context of near-native conformations of a protein. In addition, we propose an empirical site-site distance-dependent correction that can be used in conjunction with any of these models. The set of near-native structures consisted of 39 conformations of bovine pancreatic trypsin inhibitor (BPTI) each of which was a local minimum of an empirical energy function (ECEPP) in the absence of solvent. Root-mean-square (rms) deviations from the crystallographically determined structure were in the following ranges: 1.06-1.94 A for all heavy atoms, 0.77-1.36 A for all backbone heavy atoms, 0.68-1.33 A for all alpha-carbon atoms, and 1.41-2.72 A for all side-chain heavy atoms. We have found that there is considerable variation among the solvent models when evaluated in terms of concordance between the solvation free energy and the rms deviations from the crystallographically determined conformation. The solvation model for which the best concordance (0.939) with the rms deviations of the C alpha atoms was found was derived from NMR coupling constants of peptides in water combined with an exponential site-site distance dependence of the potential of mean force. Our results indicate that solvation free energy parameters derived from nonpeptide free energies of hydration may not be transferrable to peptides. Parameters derived from peptide and protein data may be more applicable to conformational analysis of proteins. A general approach to derive parameters for free energy of hydration from ensemble-averaged properties of peptides in solution is described.     

Conformations of angiotensin II and enkephalin related to N.M.R.

Statistical samples of conformations for Angiotensin II and Enkephalin, obtained from a Monte-Carlo method, are models proposed to simulate the behavior of these hormones under different physico-chemical conditions. Analysis of molecular conformations shows that, for the two peptides, extended conformations are more likely to be present in acidic solutions used for N.M.R. measurements and folded conformations due to interactions of charged terminal groups are favoured at neutral pH.     

Necessary conditions for avoiding incorrect polypeptide folds in conformational search by energy minimization

Low energy conformations have been generated for melittin, pancreatic polypeptide, and ribonuclease S-peptide, both in the vicinity of x-ray structures by energy refinement and by an unconstrained search over the entire conformational space. Since the structural polymorphism of these medium-sized peptides in crystal and solution is moderate, comparing the calculated conformations to x-ray and nmr data provides information on local and global behavior of potential functions. Local analysis includes standardization calculations, which show that models with standard geometry can approximate good resolution x-ray data with less than 0.5 Å rms deviation (RMSD). However, the atomic coordinates are shifted up to 2 Å RMSD by local energy minimization, and thus 2 Å is generally the smallest RMSD value one can target in a conformational search using the same energy evaluation models. The unconstrained search was performed by a buildup-type method based on dynamic programming. To accelerate the generation of structures in the conformational search, we used the ECEPP potential, defined in terms of standard polypeptide geometry. A number of low energy conformations were further refined by relaxing the assumption of standard bond lengths and bond angles through the use of the CHARMM potential, and the hydrophobic folding energies of Eisenberg and McLachlan were calculated. Each conformation is described in terms of the RMSD from the native, hydrogen-bonding structure, solvent-acessible surface area, and the ratio of surfaces corresponding to nonpolar and polar residues. The unconstrained search finds conformations that are different from the native, sometimes substantially, and in addition, have lower conformational energies than the native. The origin of deviations is different for each of the three peptides, but in all examples the refined x-ray structures have lower energies than the calculated incorrect folds when (1) the assumption of standard bond lengths and bond angles is relaxed; (2) a small and constant effective dielectric permittivity (ε < 10) is used; and (3) the hydrophobic folding energy is incorporated into the potential. © 1993 John Wiley & Sons, Inc.     

Crystal structures of mitochondrial processing peptidase reveal the mode for specific cleavage of import signal sequences

BACKGROUND: Mitochondrial processing peptidase (MPP) is a metalloendopeptidase that cleaves the N-terminal signal sequences of nuclear-encoded proteins targeted for transport from the cytosol to the mitochondria. Mitochondrial signal sequences vary in length and sequence, but each is cleaved at a single specific site by MPP. The cleavage sites typically contain an arginine at position -2 (in the N-terminal portion) from the scissile peptide bond in addition to other distal basic residues, and an aromatic residue at position +1. Mitochondrial import machinery recognizes amphiphilic helical conformations in signal sequences. However, it is unclear how MPP specifically recognizes diverse presequence substrates. RESULTS: The crystal structures of recombinant yeast MPP and a cleavage-deficient mutant of MPP complexed with synthetic signal peptides have been determined. MPP is a heterodimer; its alpha and beta subunits are homologous to the core II and core I proteins, respectively, of the ubiquinol-cytochrome c oxidoreductase complex. Crystal structures of two different synthetic substrate peptides cocrystallized with the mutant MPP each show the peptide bound in an extended conformation at the active site. Recognition sites for the arginine at position -2 and the +1 aromatic residue are observed. CONCLUSIONS: MPP bound two mitochondrial import presequence peptides in extended conformations in a large polar cavity. The presequence conformations differ from the amphiphilic helical conformation recognized by mitochondrial import components. Our findings suggest that the presequences adopt context-dependent conformations through mitochondrial import and processing, helical for recognition by mitochondrial import machinery and extended for cleavage by the main processing component.     

Conformational study of Ac-Xaa-Pro-NHMe dipeptides: proline puckering and trans/cis imide bond.

The conformational study on 20 Ac-Xaa-Pro-NHMe dipeptides has been carried out using an empirical potential function ECEPP/3 in order to investigate the factors responsible for the preference of proline puckering of the peptides with the trans or cis imide bond preceding the proline. The general conformational preference for down- and up-puckered dipeptides is calculated as trans-down > trans-up > cis-down > cis-up, which is reasonably in accord with that estimated by analyzing X-ray structures of proteins and the result for the single proline residue. The overestimated occurrence of trans-down conformations of proline seems to be caused by excluding long-range interactions that short dipeptides cannot have. The average computed occurrence of dipeptides with cis imide bonds is about 3%, somewhat lower than the value calculated for Ac-Pro-NHMe, which is close to experimental estimates obtained from X-ray structures of proteins. In particular, the interaction of the aromatic side chain of Xaa residue with the proline ring appears not to be strong enough to stabilize the stacked conformations of small dipeptides with cis imide bonds. The propensity to adopt trans or cis imide bond and to form secondary structures of Xaa-Pro sequences is discussed and compared with results obtained from X-ray structures of proteins.     

Observations on the gel chromatography of collagen-like peptides and use of a simple calibration method for molecular weight determination.

Elution characteristics of collagen-derived polypeptides and of globular proteins were compared under identical experimental conditions with agarose gels. This comparison permitted calculation of the hydrodynamic radii of collagen polypeptide chains of different molecular weight, and these radii were shown to be in reasonable agreement with estimates made from intrinsic viscosity data. Two distinet linear relationships were observed for collagen polypeptide chains, relating logarithm of molecular weight to elution parameters. Peptide chains of MW 3300 and lower fell on a line of a steeper slope than did larger polypeptide chains.A simple procedure for molecular weight estimation of an unknown polypeptide chain of the collagen class is described, using only three commercially available standards for calibration: reduced, carboxymethylated Ascaris cuticle collagen, and the synthetic peptides (l-Pro-l-Pro-Gly)₁₀ and (l-Pro-L-Pro-Gly)₅.     

Conformational preferences and the role of the statine residue in the crystal state

The present paper is the result of an analysis of the available crystal structure data related to the statine amino acid so as to obtain information about bond lengths, bond angles, and preferential conformations. The number of configurations actually observed is small; nevertheless, some characteristic conformations should be pointed out for statine-containing peptides. The presence of two additional carbon atoms in the statine main chain enhances the peptide conformational degree of freedom and the statine-containing peptides are observed in a variety of different conformations including some usual secondary structure-types as beta-turns and beta-strands.     

Enhanced stability of cis Pro-Pro peptide bond in Pro-Pro-Phe sequence motif

Identification of sequence motifs that favor cis peptide bonds in proteins is important for understanding and designing proteins containing turns mediated by cis peptide conformations. From (1)H NMR solution studies on short peptides, we show that the Pro-Pro peptide bond in Pro-Pro-Phe almost equally populates the cis and trans isomers, with the cis isomer stabilized by a CHc...pi interaction involving the terminal Pro and Phe. We also show that Phe is over-represented at sequence positions immediately following cis Pro-Pro motifs in known protein structures. Our results demonstrate that the Pro-Pro cis conformer in Pro-Pro-Phe sequence motifs is as important as the trans conformer, both in short peptides as well as in natively folded proteins.     

Periodic conformations of deoxyribonucleic acids

The conformation of a deoxyribonucleotide unit in a deoxyribonucleic acid molecule can be defined by six angles, each of which specifies the relative orientations between two groups of atoms adjacent to a covalent bond. With the assumption that these atoms are hard spheres with fixed van der Waal radii, conformations are sought to minimize their overlapping. The additional requirement of the polymer having a periodic structure further reduces the allowable conformations.     

The volume of cavities in proteins and virus capsids

An improved algorithm for the calculation of the volume of internal cavities within protein structures and virus capsids as well as the volumes occupied by single amino acid residues were presented. The geometrical approach was based on atomic van der Waals radii. The results obtained with two sets of the radii, those proposed by Pauling and those determined by Tsai et al were compared. The main improvement compared with our previous approach is a more elaborate treatment of the regions at the very boundary of the cavities, which yields a more accurate volume estimate. The cavity volume of a number of Plant Pathogenesis‐Related proteins of class 10 (PR‐10) were reevaluated and the volumes and other geometrical parameters for about 400 capsids of icosahedral viruses were reported. Using the same approach the volumes of amino acid residues in polypeptides as mean values averaged over multiple conformations of the side chain were also estimated. Proteins 2016; 84:1275–1286. © 2016 Wiley Periodicals, Inc.