The link between sequence and conformation in protein structures appears to be stereochemically established

In search of the link between sequence and conformation in protein structures, we perform molecular dynamics analysis of the effect of stereochemical mutation in end-protected octa-alanine Ac-Ala8-NHMe from poly-L to an alternating-L,D structure. The mutation has a dramatic effect, transforming the peptide from a condition of extreme sensitivity to one of extreme insensitivity to solvent. Examining the molecular folds of poly-L and alternating-L,D structure in atomistic detail, we find them to differ in the relationship between peptide dipolar interactions at the local and nonlocal levels, either conflicting or harmonious depending upon the chain stereochemistry. The stereochemical transformation of interpeptide electrostatics from a condition of conflict to one of harmony explains the long-standing puzzle of why poly-L and alternating-L,D peptides strongly differ in properties such as "stiffness" and solvent sensitivity. Furthermore, it is possible that poly-L stereochemistry is also the fulcrum of protein sensitivity to the effects of amino acid side-chain structures via dielectric arbitrations in interpeptide electrostatics. Indeed the evidence is accumulating that the amino acid side chains differing in alpha-helix and beta-sheet propensities also differ in their desolvating effects in the adjacent and nearest-neighbor peptides and thus possibly in the solvent screening of peptide dipolar interactions.     

Intrinsic folding of small peptide chains: spectroscopic evidence for the formation of beta-turns in the gas phase

Laser desorption of model peptides coupled to laser spectroscopic techniques enables the gas-phase observation of genuine secondary structures of biology. Spectroscopic evidence for the formation of beta-turns in gas-phase peptide chains containing glycine and phenylalanine residues establishes the intrinsic stability of these forms and their ability to compete with other stable structures. The precise characterization of local minima on the potential energy surface from IR spectroscopy constitutes an acute assessment for the state-of-the-art quantum mechanical calculations also presented. The observation of different types of beta-turns depending upon the residue order within the sequence is found to be consistent with the residue propensities in beta-turns of proteins, which suggests that the prevalence of glycine in type II and II' turns stems essentially from an energetic origin, already at play under isolated conditions.     

Identification of potential small molecule peptidomimetics similar to motifs in proteins

Protein-protein interactions are central to most biological processes and represent a large and important class of targets for human therapeutics. Small molecules containing peptide substituents may mimic regions of interacting proteins and inhibit their interactions. We set out to develop efficient methods to screen for similarities between known peptide structures within proteins and small molecules. We developed a method to rank peptide-compound similarities, that is restricted to small linear motifs in proteins, and to compounds containing amino acid substituents. Application to a search of the PubChem database (5.4 million compounds) using all short motifs on accessible surface areas in a nonredundant set of 11 488 peptides from the protein structure database PDB demonstrated the feasibility of the method for high throughput comparisons and the availability of compounds with comparable substituents: over 6 million compound-peptide pairs shared at least three amino acid substituents, approximately 100 000 of which had an rmsd score of less than 1 A. A Z-score function was developed that compares matches of a compound to different instances of the peptide motif in PDB, providing an appropriate scoring function for comparison among peptide-compound similarities involving different numbers of atoms (while simultaneously enriching for similarities that are likely to be more specific for the protein of interest). We applied the method to searches of known short protein motifs against the National Cancer Institute Developmental Therapeutic Program compound database, identifying a known true positive.     

Microwave-assisted acid and base hydrolysis of intact proteins containing disulfide bonds for protein sequence analysis by mass spectrometry

Controlled hydrolysis of proteins to generate peptide ladders combined with mass spectrometric analysis of the resultant peptides can be used for protein sequencing. In this paper, two methods of improving the microwave-assisted protein hydrolysis process are described to enable rapid sequencing of proteins containing disulfide bonds and increase sequence coverage, respectively. It was demonstrated that proteins containing disulfide bonds could be sequenced by MS analysis by first performing hydrolysis for less than 2 min, followed by 1 h of reduction to release the peptides originally linked by disulfide bonds. It was shown that a strong base could be used as a catalyst for microwave-assisted protein hydrolysis, producing complementary sequence information to that generated by microwave-assisted acid hydrolysis. However, using either acid or base hydrolysis, amide bond breakages in small regions of the polypeptide chains of the model proteins (e.g., cytochrome c and lysozyme) were not detected. Dynamic light scattering measurement of the proteins solubilized in an acid or base indicated that protein-protein interaction or aggregation was not the cause of the failure to hydrolyze certain amide bonds. It was speculated that there were some unknown local structures that might play a role in preventing an acid or base from reacting with the peptide bonds therein.     

Dynamic Amino Acid Side-Chains Grafting on Folded Peptide Backbone**

An efficient strategy for the synthesis of large libraries of conformationally defined peptides is reported, using dynamic combinatorial chemistry as a tool to graft amino acid side chains on a well-ordered 3D (3-dimension) peptide backbone. Combining rationally designed scaffolds with combinatorial side chains selection represents an alternative method to access peptide libraries for structures that are not genetically encodable. This method would allow a breakthrough for the discovery of protein mimetic for unconventional targets for which little is known.     

多肽序列的结构特征与其MHC限制性

从多肽序列的一级结构出发,基于多肽序列中氨基酸侧链间距离和氨基酸侧链的电性特征,构建了多肽序列特征矢量,简称ζ矢量,选取文献中主要组织相容性复合物(MHC)中的14个A^d限制性和14个非A^d限制性多肽序列作为训练集建立辅助性T细胞(helperTlymphocytc,Th)表位预测的定量模型,为了检验该预报系统的精确性,进行了随机抽样检验和交互检验.结果表明,该预报系统具有稳定性好、预测能力强的特点,该方法可用于人的MHCⅠ和Ⅱ类表位的预测、蛋白质抗原免疫识别、亚单位疫苗分子设计及研制.     

Efficient molecular mechanics simulations of the folding,orientation, and assembly of peptides in lipid bilayers using an implicit atomic solvation model

Membrane proteins comprise a significant fraction of the proteomes of sequenced organisms and are the targets of approximately half of marketed drugs. However, in spite of their prevalence and biomedical importance, relatively few experimental structures are available due to technical challenges. Computational simulations can potentially address this deficit by providing structural models of membrane proteins. Solvation within the spatially heterogeneous membrane/solvent environment provides a major component of the energetics driving protein folding and association within the membrane. We have developed an implicit solvation model for membranes that is both computationally efficient and accurate enough to enable molecular mechanics predictions for the folding and association of peptides within the membrane. We derived the new atomic solvation model parameters using an unbiased fitting procedure to experimental data and have applied it to diverse problems in order to test its accuracy and to gain insight into membrane protein folding. First, we predicted the positions and orientations of peptides and complexes within the lipid bilayer and compared the simulation results with solid-state NMR structures. Additionally, we performed folding simulations for a series of host–guest peptides with varying propensities to form alpha helices in a hydrophobic environment and compared the structures with experimental measurements. We were also able to successfully predict the structures of amphipathic peptides as well as the structures for dimeric complexes of short hexapeptides that have experimentally characterized propensities to form beta sheets within the membrane. Finally, we compared calculated relative transfer energies with data from experiments measuring the effects of mutations on the free energies of translocon-mediated insertion of proteins into lipid bilayers and of combined folding and membrane insertion of a beta barrel protein.     

Constraining Cyclic Peptides To Mimic Protein Structure Motifs

Many proteins exert their biological activities through small exposed surface regions called epitopes that are folded peptides of well‐defined three‐dimensional structures. Short synthetic peptide sequences corresponding to these bioactive protein surfaces do not form thermodynamically stable protein‐like structures in water. However, short peptides can be induced to fold into protein‐like bioactive conformations (strands, helices, turns) by cyclization, in conjunction with the use of other molecular constraints, that helps to fine‐tune three‐dimensional structure. Such constrained cyclic peptides can have protein‐like biological activities and potencies, enabling their uses as biological probes and leads to therapeutics, diagnostics and vaccines. This Review highlights examples of cyclic peptides that mimic three‐dimensional structures of strand, turn or helical segments of peptides and proteins, and identifies some additional restraints incorporated into natural product cyclic peptides and synthetic macrocyclic peptidomimetics that refine peptide structure and confer biological properties.     

Highly ordered structures of peptides by using molecular scaffolds

Protein secondary structures such as alpha-helices, beta-sheets, and beta-turns are important in inducing the three-dimensional structure and biological activity of proteins. Designing secondary structure mimics composed of short peptides has attracted much attention not only to gain fundamental insight into the factors affecting protein folding but also to develop pharmacologically useful compounds, artificial receptors, asymmetric catalysts, and new materials. In this tutorial review, we focus on molecular scaffolds employed to induce beta-sheet-like structure in attached peptide chains, thereby creating highly ordered molecular structures, and discuss the versatility of these molecular scaffolds to regulate the attached peptide strands in the appropriate dimensions.     

How sensitive are nanosecond molecular dynamics simulations of proteins to changes in the force field?

The sensitivity of molecular dynamics simulations to variations in the force field has been examined in relation to a set of 36 structures corresponding to 31 proteins simulated by using different versions of the GROMOS force field. The three parameter sets used (43a1, 53a5, and 53a6) differ significantly in regard to the nonbonded parameters for polar functional groups and their ability to reproduce the correct solvation and partitioning behavior of small molecular analogues of the amino acid side chains. Despite the differences in the force field parameters no major differences could be detected in a wide range of structural properties such as the root-mean-square deviation from the experimental structure, radii of gyration, solvent accessible surface, secondary structure, or hydrogen bond propensities on a 5 to 10 ns time scale. The small differences that were observed correlated primarily with the presence of charged residues as opposed to residues that differed most between the parameter sets. The work highlights the variation that can be observed in nanosecond simulations of protein systems and implications of this for force field validation, as well as for the analysis of protein simulations in general.     

Structural dependencies of h3JNC' scalar coupling in protein H-bond chains

The H-bond ((h3)J(NC')) and peptide bond ((1)J(NC')) scalar couplings establish connectivity of the electronic structure in the H-bond chains of proteins. The correlated changes of (h3)J(NC') and (1)J(NC') couplings extend over several peptide groups in the chains. Consequently, the electronic structure of the H-bond chains can affect (h3)J(NC') in a manner that is independent of the local H-bond geometry. By taking this into account, and by using a more complete set of H-bond geometry parameters, we have predicted (h3)J(NC') couplings in the H-bond chains with deviations commensurate to the standard deviations of the experimentally determined values. We have created a comprehensive database of (h3)J(NC') and (1)J(NC') couplings by measuring the coupling constants in ubiquitin (alphabeta-fold) intestinal fatty acid binding protein (beta-barrel) and carp parvalbumin (alpha-helical).     

Stability of cyclic beta-hairpins: asymmetric contributions from side chains of a hydrogen-bonded cross-strand residue pair

Amino acid structural propensities measured in "host-guest" model studies are often used in protein structure prediction or to choose appropriate residues in de novo protein design. While this concept has proven useful for helical structures, it is more difficult to apply successfully to beta-sheets. We have developed a cyclic beta-hairpin scaffold as a host for measurement of individual residue contributions to hairpin structural stability. Previously, we have characterized substitutions in non-backbone-hydrogen-bonded strand sites; relative stability differences measured in the cyclic host are highly predictive of changes in folding free energy for linear beta-hairpin peptides. Here, we examine the hydrogen-bonded strand positions of our host. Surprisingly, we find a large favorable contribution to stability from a valine (or isoleucine) substitution immediately preceding the C-terminal cysteine of the host peptide, but not at the cross-strand position of the host or in either strand of a folded linear beta-hairpin (trpzip peptide). Further substitutions in the peptides and NMR structural analysis indicate that the stabilizing effect of valine is general for CX(8)C cyclic hairpins and cannot be explained by particular side-chain-side-chain interactions. Instead, a localized decrease in twist of the peptide backbone on the N-terminal side of the cysteine allows the valine side chain to adopt a unique conformation that decreases the solvent accessibility of the peptide backbone. The conformation differs from the highly twisted (coiled) conformation of the trpzip hairpins and is more typical of conformations present in multistranded beta-sheets. This unexpected structural fine-tuning may explain why cyclic hairpins selected from phage-displayed libraries often have valine in the same position, preceding the C-terminal cysteine. It also emphasizes the diversity of structures accessible to beta-strands and the importance of considering not only "beta-propensity", but also hydrogen-bonding pattern and strand twist, when designing beta structures. Finally, we observe correlated, cooperative stabilization from side-chain substitutions on opposite faces of the hairpin. This suggests that cooperative folding in beta-hairpins and other small beta-structures is driven by cooperative strand-strand association.     

A Multicomponent Conjugation Strategy to Unique N‐Steroidal Peptides: First Evidence of the Steroidal Nucleus as a β‐Turn Inducer in Acyclic Peptides

Constraining small peptides into specific secondary structures has been a major challenge in peptide ligand design. So far, the major solution for decreasing the conformational flexibility in small peptides has been cyclization. An alternative is the use of topological templates, which are able to induce and/or stabilize peptide secondary structures by means of covalent attachment to the peptide. Herein a multicomponent strategy and structural analysis of a new type of peptidosteroid architecture having the steroid as N‐substituent of an internal amide bond is reported. The approach comprises the one‐pot conjugation of two peptide chains (or amino acid derivatives) to aminosteroids by means of the Ugi reaction to give a unique family of N‐steroidal peptides. The conjugation efficiency of a variety of peptide sequences and steroidal amines, as well as their consecutive head‐to‐tail cyclization to produce chimeric cyclopeptide–steroid conjugates, that is, macrocyclic lipopeptides, was assessed. Determination of the three‐dimensional structure of an acyclic N‐steroidal peptide in solution proved that the bulky, rigid steroidal template is capable of both increasing significantly the conformational rigidity, even in a peptide sequence as short as five amino acid residues, and inducing a β‐turn secondary structure even in the all‐s‐trans isomer. This report provides the first evidence of the steroid skeleton as β‐turn inducer in linear peptide sequences.     

Dipole-induced self-assembly of helical beta-peptides

In this work, the interactions between beta-peptides are investigated for helix-forming peptides using molecular simulation. The role of electrostatic interactions in the self-assembly of these peptides is studied by calculating the dipole moment of various 14-helical beta-peptides using molecular dynamics simulations. The stability of a beta-peptide that is known to form a liquid crystalline phase is determined by calculating the potential of mean force using the expanded ensemble density of states method. This peptide is found to form a mechanically stable 14-helix in an implicit solvent model. The interaction between two of these peptides is examined by calculating the potential of mean force between the two peptides in implicit solvent. The peptides are shown to favorably associate in an end-to-end manner, driven largely by dipolar interactions. In order to understand the possible structures that form when many peptides interact in solution, a coarse-grained model is developed. Brownian dynamics simulations of the coarse-grained model at intermediate concentrations (1-50 mM) are performed, and the aggregation behavior is understood by calculating the diffusivity and the radial distribution function. An analysis of the resultant structures reveals that the coarse-grained model of the peptide leads to the formation of spherical clusters.     

Peptide/protein-polymer conjugates: synthetic strategies and design concepts

This feature article provides a compilation of tools available for preparing well-defined peptide/protein-polymer conjugates, which are defined as hybrid constructs combining (i) a defined number of peptide/protein segments with uniform chain lengths and defined monomer sequences (primary structure) with (ii) a defined number of synthetic polymer chains. The first section describes methods for post-translational, or direct, introduction of chemoselective handles onto natural or synthetic peptides/proteins. Addressed topics include the residue- and/or site-specific modification of peptides/proteins at Arg, Asp, Cys, Gln, Glu, Gly, His, Lys, Met, Phe, Ser, Thr, Trp, Tyr and Val residues and methods for producing peptides/proteins containing non-canonical amino acids by peptide synthesis and protein engineering. In the second section, methods for introducing chemoselective groups onto the side-chain or chain-end of synthetic polymers produced by radical, anionic, cationic, metathesis and ring-opening polymerization are described. The final section discusses convergent and divergent strategies for covalently assembling polymers and peptides/proteins. An overview of the use of chemoselective reactions such as Heck, Sonogashira and Suzuki coupling, Diels-Alder cycloaddition, Click chemistry, Staudinger ligation, Michael's addition, reductive alkylation and oxime/hydrazone chemistry for the convergent synthesis of peptide/protein-polymer conjugates is given. Divergent approaches for preparing peptide/protein-polymer conjugates which are discussed include peptide synthesis from synthetic polymer supports, polymerization from peptide/protein macroinitiators or chain transfer agents and the polymerization of peptide side-chain monomers.     

Construction of a protein-detection system using a loop peptide library with a fluorescence label

Construction of a novel protein-detection system was carried out using a designed peptide library with fluorescent labels based on loop structures. As a basic model study, detection of alpha-amylase using fluorescent-labeled peptides derived from an active loop of tendamistat was examined. The detection methods for proteins with immobilized peptides as well as peptides in solution have been successfully established. Based on these results, a loop peptide library that has various turn sequences grafted on a stable loop structure has been constructed. Various proteins with recognition patterns corresponding, for instance, to "protein fingerprints" could be detected using an immobilized peptide library. The present results suggest that the system can be applied to the development of a peptide microarray that behaves as a protein chip.     

The Impact of Amino Acid Side Chain Mutations in Conformational Design of Peptides and Proteins

Local energetic effects of amino acid replacements are often considered to have only a moderate influence on the backbone conformation of proteins or peptides. As these effects are difficult to determine experimentally, no comparison has yet been performed. However, knowledge of the influence of side chain mutations is essential in protein homology modeling and in optimizing biologically active peptide ligands in medicinal chemistry. Furthermore, the tool of N‐methylation of peptides is of increasing importance for the design of peptidic drugs to gain oral availability or receptor selectivity. However, N‐methylation is often accompanied by considerable population of cis‐peptide bond structures, resulting in completely different conformations compared with the parent peptide. To retain a favored structure, it might be important to understand the effect of different side chains on the backbone conformation and to enable the introduction of an N‐methylation at the right position without disturbing a biologically active conformation. In order to detect even small energetic effects due to side chain mutations, we employed a trick to investigate the structural equilibrium of a selected cyclic pentapeptide in which two conformations are equally populated. Very small energetic differences between both conformations could easily be determined experimentally by identifying shifts in the population of both isomers.     

Structural regulation of a peptide-conjugated graft copolymer: a simple model for amyloid formation

The self-assembly of peptides and proteins into beta-sheet-rich high-order structures has attracted much attention as a result of the characteristic nanostructure of these assemblies and because of their association with neurodegenerative diseases. Here we report the structural and conformational properties of a peptide-conjugated graft copolymer, poly(gamma-methyl-L-glutamate) grafted polyallylamine (1) in a water-2,2,2-trifluoroethanol solution as a simple model for amyloid formation. Atomic force microscopy revealed that the globular peptide 1 self-assembles into nonbranching fibrils that are about 4 nm in height under certain conditions. These fibrils are rich in beta-sheets and, similar to authentic amyloid fibrils, bind the amyloidophilic dye Congo red. The secondary and quaternary structures of the peptide 1 can be controlled by manipulating the pH, solution composition, and salt concentration; this indicates that the three-dimensional packing arrangement of peptide chains is the key factor for such fibril formation. Furthermore, the addition of carboxylic acid-terminated poly(ethylene glycol), which interacts with both of amino groups of 1 and hydrophobic PMLG chains, was found to obviously inhibit the alpha-to-beta structural transition for non-assembled peptide 1 and to partially cause a beta-to-alpha structural transition against the 1-assembly in the beta-sheet form. These findings demonstrate that the amyloid fibril formation is not restricted to specific protein sequences but rather is a generic property of peptides. The ability to control the assembled structure of the peptide should provide useful information not only for understanding the amyloid fibril formation, but also for developing novel peptide-based material with well-defined nanostructures.     

Aromatic-aromatic interactions in crystal structures of helical peptide scaffolds containing projecting phenylalanine residues

Aromatic-aromatic interactions between phenylalanine side chains in peptides have been probed by the structure determination in crystals of three peptides: Boc-Val-Ala-Phe-Aib-Val-Ala-Phe-Aib-OMe, I; Boc-Val-Ala-Phe-Aib-Val-Ala-Phe-Aib-Val-Ala-Phe-Aib-OMe, II; Boc-Aib-Ala-Phe-Aib-Phe-Ala-Val-Aib-OMe, III. X-ray diffraction studies reveal that all three peptides adopt helical conformations in the solid state with the Phe side chains projecting outward. Interhelix association in the crystals is promoted by Phe-Phe interactions. A total of 15 unique aromatic pairs have been characterized in the three independent crystal structures. In peptides I and II, the aromatic side chains lie on the same face of the helix at i/i + 4 positions resulting in both intrahelix and interhelix aromatic interactions. In peptide III, the Phe side chains are placed on the opposite faces of the helix, resulting in exclusive intermolecular aromatic interactions. The distances between the centroids of aromatic pair ranges from 5.11 to 6.86 A, while the distance of closest approach of ring carbon atoms ranges from 3.27 to 4.59 A. Examples of T-shaped and parallel-displaced arrangements of aromatic pairs are observed, in addition to several examples of inclined arrangements. The results support the view that the interaction potential for a pair of aromatic rings is relatively broad and rugged with several minima of similar energies, separated by small activation barriers.