Peptoid oligomers with alpha-chiral, aromatic side chains: sequence requirements for the formation of stable peptoid helices.

The achiral backbone of oligo-N-substituted glycines or "peptoids" lacks hydrogen-bond donors, effectively preventing formation of the regular, intrachain hydrogen bonds that stabilize peptide alpha-helical structures. Yet, when peptoids are N-substituted with alpha-chiral, aromatic side chains, oligomers with as few as five residues form stable, chiral, polyproline-like helices in either organic or aqueous solution. The adoption of chiral secondary structure in peptoid oligomers is primarily driven by the steric influence of these bulky, chiral side chains. Interestingly, peptoid helices of this class exhibit intense circular dichroism (CD) spectra that closely resemble those of peptide alpha-helices. Here, we have taken advantage of this distinctive spectroscopic signature to investigate sequence-related factors that favor and disfavor stable formation of peptoid helices of this class, through a comparison of more than 30 different heterooligomers with mixed chiral and achiral side chains. For this family of peptoids, we observe that a composition of at least 50% alpha-chiral, aromatic residues is necessary for the formation of stable helical structure in hexameric sequences. Moreover, both CD and 1H-13C HSQC NMR studies reveal that these short peptoid helices are stabilized by the placement of an alpha-chiral, aromatic residue on the carboxy terminus. Additional stabilization can be provided by the presence of an "aromatic face" on the helix, which can be patterned by positioning aromatic residues with three-fold periodicity in the sequence. Extending heterooligomer chain length beyond 12-15 residues minimizes the impact of the placement, but not the percentage, of alpha-chiral aromatic side chains on overall helical stability. In light of these new data, we discuss implications for the design of helical, biomimetic peptoids based on this structural motif.     

SPINE X: improving protein secondary structure prediction by multistep learning coupled with prediction of solvent accessible surface area and backbone torsion angles

Accurate prediction of protein secondary structure is essential for accurate sequence alignment, three-dimensional structure modeling, and function prediction. The accuracy of ab initio secondary structure prediction from sequence, however, has only increased from around 77 to 80% over the past decade. Here, we developed a multistep neural-network algorithm by coupling secondary structure prediction with prediction of solvent accessibility and backbone torsion angles in an iterative manner. Our method called SPINE X was applied to a dataset of 2640 proteins (25% sequence identity cutoff) previously built for the first version of SPINE and achieved a 82.0% accuracy based on 10-fold cross validation (Q(3)). Surpassing 81% accuracy by SPINE X is further confirmed by employing an independently built test dataset of 1833 protein chains, a recently built dataset of 1975 proteins and 117 CASP 9 targets (critical assessment of structure prediction techniques) with an accuracy of 81.3%, 82.3% and 81.8%, respectively. The prediction accuracy is further improved to 83.8% for the dataset of 2640 proteins if the DSSP assignment used above is replaced by a more consistent consensus secondary structure assignment method. Comparison to the popular PSIPRED and CASP-winning structure-prediction techniques is made. SPINE X predicts number of helices and sheets correctly for 21.0% of 1833 proteins, compared to 17.6% by PSIPRED. It further shows that SPINE X consistently makes more accurate prediction in helical residues (6%) without over prediction while PSIPRED makes more accurate prediction in coil residues (3-5%) and over predicts them by 7%. SPINE X Server and its training/test datasets are available at http://sparks.informatics.iupui.edu/     

Improved helix and kink characterization in membrane proteins allows evaluation of kink sequence predictors

Although the α-helical secondary structure of proteins is well-defined, the exact causes and structures of helical kinks are not. This is especially important for transmembrane (TM) helices of integral membrane proteins, many of which contain kinks providing functional diversity despite predominantly helical structure. We have developed a Monte Carlo method based algorithm, MC-HELAN, to determine helical axes alongside positions and angles of helical kinks. Analysis of all nonredundant high-resolution α-helical membrane protein structures (842 TM helices from 205 polypeptide chains) revealed kinks in 64% of TM helices, demonstrating that a significantly greater proportion of TM helices are kinked than those indicated by previous analyses. The residue proline is over-represented by a factor >5 if it is two or three residues C-terminal to a bend. Prolines also cause kinks with larger kink angles than other residues. However, only 33% of TM kinks are in proximity to a proline. Machine learning techniques were used to test for sequence-based predictors of kinks. Although kinks are somewhat predicted by sequence, kink formation appears to be driven predominantly by other factors. This study provides an improved view of the prevalence and architecture of kinks in helical membrane proteins and highlights the fundamental inaccuracy of the typical topological depiction of helical membrane proteins as series of ideal helices.     

A general treatment of the configuration statistics of polysaccharides.

A treatment of the configurational statistics of polysaccharides is given in the isomeric state approximation. All classes of linear polysaccharides of specifiec chemical sequence are treated simultaneously. Chain tortuosity arising from torsional motions about the chemical bonds of the glycosidic linkages is recognized explicitly as is the possibility for conformational isomerism of the sugar residues. Valence angles and lengths are taken to be fixed at the equilibrium values, and pyranose residues in their chair conformations are treated as inflexible constituents of the skeletal structure. Pyranose and furanose forms capable of pseudorotation may be incorporated as rigid skeletal entities as well, provided suitable attention is given to the selection and interpretation of the conformational isomeric states included. Separation of the configuration energy into independent contributions is shown to be impossible in general. Methods are described for assessing the influence of neighbor interactions on the populations of the several conformers of the sugar residues. The relative conformational free energy of the flexible and chair form conformers of pyranose sugars is discussed, and appropriate measures of polysaccharide chain flexibility and stiffness are suggested.     

Formation of Periodic γ‐Turns in α/β‐Hybrid Peptides: DFT and NMR Experimental Evidence

Hybrid peptidic oligomers comprising natural and unnatural amino acid residues that can exhibit biomolecular folding and hydrogen‐bonding mimicry have attracted considerable interest in recent years. While a variety of hybrid peptidic helices have been reported in the literature, other secondary structural patterns such as γ‐turns and ribbons have not been well explored so far. The present work reports the design of novel periodic γ‐turns in the oligomers of 1:1 natural‐α/unnatural trans‐β‐norborenene (TNAA) amino acid residues. Through DFT, NMR, and MD studies, it is convincingly shown that, in the mixed conformational pool, the heterogeneous backbone of the hybrid peptides preferentially adopt periodic 8‐membered (pseudo γ‐turn)/7‐membered (inverse γ‐turn) hydrogen bonds in both polar and non‐polar solvent media. It is observed that the stereochemistry and local conformational preference of the β‐amino acid building blocks have a profound influence on accessing the specific secondary fold. These findings may be of significant relevance for the development of molecular scaffolds that facilitate desired positioning of functional side‐chains.     

Peptoid oligomers with alpha-chiral, aromatic side chains: effects of chain length on secondary structure.

Oligomeric N-substituted glycines or "peptoids" with alpha-chiral, aromatic side chains can adopt stable helices in organic or aqueous solution, despite their lack of backbone chirality and their inability to form intrachain hydrogen bonds. Helical ordering appears to be stabilized by avoidance of steric clash as well as by electrostatic repulsion between backbone carbonyls and pi clouds of aromatic rings in the side chains. Interestingly, these peptoid helices exhibit intense circular dichroism (CD) spectra that closely resemble those of peptide alpha-helices. Here, we have utilized CD to systematically study the effects of oligomer length, concentration, and temperature on the chiral secondary structure of organosoluble peptoid homooligomers ranging from 3 to 20 (R)-N-(1-phenylethyl)glycine (Nrpe) monomers in length. We find that a striking evolution in CD spectral features occurs for Nrpe oligomers between 4 and 12 residues in length, which we attribute to a chain length-dependent population of alternate structured conformers having cis versus trans amide bonds. No significant changes are observed in CD spectra of oligomers between 13 and 20 monomers in length, suggesting a minimal chain length of about 13 residues for the formation of stable poly(Nrpe) helices. Moreover, no dependence of circular dichroism on concentration is observed for an Nrpe hexamer, providing evidence that these helices remain monomeric in solution. In light of these new data, we discuss chain length-related factors that stabilize organosoluble peptoid helices of this class, which are important for the design of helical, biomimetic peptoids sharing this structural motif.     

Structure of the fully modified left-handed cyclohexene nucleic acid sequence GTGTACAC

CeNA oligonucleotides consist of a phosphorylated backbone where the deoxyribose sugars are replaced by cyclohexene moieties. The X-ray structure determination and analysis of a fully modified octamer sequence GTGTACAC, which is the first crystal structure of a carbocyclic-based nucleic acid, is presented. This particular sequence was built with left-handed building blocks and crystallizes as a left-handed double helix. The helix can be characterized as belonging to the (mirrored) A-type family. Crystallographic data were processed up to 1.53 A, and the octamer sequence crystallizes in the space group R32. The sugar puckering is found to adopt the 3H2 half-chair conformation which mimics the C3'-endo conformation of the ribose sugar. The double helices stack on top of each other to form continuous helices, and static disorder is observed due to this end-to-end stacking.     

Selective covalent capture of collagen triple helices with a minimal protecting group strategy

Collagens and their most characteristic structural unit, the triple helix, play many critical roles in living systems which drive interest in preparing mimics of them. However, application of collagen mimetic helices is limited by poor thermal stability, slow rates of folding and poor equilibrium between monomer and trimer. Covalent capture of the self-assembled triple helix can solve these problems while preserving the native three-dimensional structure critical for biological function. Covalent capture takes advantage of strategically placed lysine and glutamate (or aspartate) residues which form stabilizing charge–pair interactions in the supramolecular helix and can subsequently be converted to isopeptide amide bonds under folded, aqueous conditions. While covalent capture is powerful, charge paired residues are frequently found in natural sequences which must be preserved to maintain biological function. Here we describe a minimal protecting group strategy to allow selective covalent capture of specific charge paired residues which leaves other charged residues unaltered. We investigate a series of side chain protecting groups for lysine and glutamate in model peptides for their ability to be deprotected easily and in high yield while maintaining (1) the solubility of the peptides in water, (2) the self-assembly and stability of the triple helix, and (3) the ability to covalently capture unprotected charge pairs. Optimized conditions are then illustrated in peptides derived from Pulmonary Surfactant protein A (SP-A). These covalently captured SP-A triple helices are found to have dramatically improved rates of folding and thermal stability while maintaining unmodified lysine–glutamate pairs in addition to other unmodified chemical functionality. The approach we illustrate allows for the covalent capture of collagen-like triple helices with virtually any sequence, composition or register. This dramatically broadens the utility of the covalent capture approach to the stabilization of biomimetic triple helices and thus also improves the utility of biomimetic collagens generally.

A minimal protecting group strategy is developed to allow selective covalent capture of collagen-like triple helices. This allows stabilization of this critical fold while preserving charge–pair interactions critical for biological applications.     

甘草多糖螺旋结构的原子力显微镜研究

用原子力显微镜(AFM)对甘草多糖的微观结构进行观察, 实验结果表明, 甘草多糖主要由葡萄糖、阿拉伯糖和半乳糖组成. 甘草多糖分子的稀溶液铺展在Ni^2＋处理的云母片上, 经干燥, 乙醇固定后, 获得稳定、重复的图像. 甘草多糖分子具有高度分枝的结构, 并且糖链间形成环状、柱状或近似于螺旋状的结构. 甘草多糖链呈多股紧密的螺旋结构, 这种现象可能与该多糖中分子间的Van der Waals相互作用以及糖链间氢键缔合有关.     

Conformational flexibility of the group B meningococcal polysaccharide in solution

To elucidate the role of secondary structure in the immune response against alpha(2-->8)-linked polysialic acid, the capsular polysaccharide of Group B meningococci, we have investigated its solution dynamics by using specific models of molecular motion and hydrodynamic modeling to interpret experimental NMR data. (13)C-[(1)H] NMR relaxation times and steady-state NOE enhancements were measured for two aqueous solutions of alpha(2-->8)-linked sialic acid polysaccharides. Each contained a unique distribution of polysaccharide chain lengths, with average lengths estimated at 40 or 400 residues. Models for rigid molecule tumbling, including two based on helical conformations proposed for the polysaccharide,(31) could not explain the NMR measurements. In general for these helices, the correlation times for their overall tumbling that best account for the NMR data correspond to polysaccharide chains between 9 and 18 residues in length, far short of the average lengths estimated for either solution. The effects of internal motions incorporated into these helices was modeled with an effective correlation time representing helix tumbling as well as internal motion. This modeling demonstrated that even with extreme amounts of internal motion, "flexible helices" of 25 residues or more still could not produce the NMR measurements. All data are consistent with internal and segmental motions dominating the nuclear magnetic relaxation of the polysaccharide and not molecular tumbling. Statistical distributions of correlation times have been found specifically for the pyranose rings, linkage groups, and methoxy groups that can account for the measured relaxation times and NOE enhancements. The distributions suggest that considerable flexibility attends the polysaccharide in solution, and the ranges of motional frequencies for the linkage groups and pyranose rings are comparable. We conclude that the Group B meningococcal polysaccharide is a random coil chain in solution, and therefore, does not have antigenic epitopes dependent upon a rigid, ordered conformation.     

Controlling the morphology of cross beta-sheet assemblies by rational design

Low molecular weight peptidomimetics with simple amphiphilic sequences can help to elucidate the structures of cross beta-sheet assemblies, such as amyloid fibrils. The peptidomimetics described herein comprise a dibenzofuran template, two peptide strands made up of alternating hydrophilic and hydrophobic residues, and carboxyl termini, each of which can be varied to probe the structural requirements for beta-sheet self-assembly processes. The dibenzofuran template positions the strands approximately 10 A apart, allowing corresponding hydrophobic side chains in the strands to pack into a collapsed U-shaped structure. This conformation is stabilized by hydrophobic interactions, not intramolecular hydrogen bonds. Intermolecular stacking of the collapsed peptidomimetics, enabled by intermolecular hydrogen bonding and hydrophobic interactions, affords 25-27 A wide protofilaments having a cross beta-sheet structure. Association of protofilaments, mediated by the dibenzofuran substructures and driven by the hydrophobic effect, affords 50-60 A wide filaments. These widths can be controlled by changing the length of the peptide strands. Further assembly of the filaments into fibrils or ribbons can be controlled by modification of the template, C-terminus, and buffer ion composition.     

Characterization of alpha helices interacting with nucleic acids

Protein-nucleic acid interactions play a vital role in most genetic processes. An enhanced insight into such interactions can be obtained from the structure database of these complexes. Here, we report an overall survey on the geometry of alpha helices which interact with nucleic acids through hydrogen bonds and/or non-bonded interactions. Using the program RADIL based on an algorithm developed from this laboratory, 161 alpha helices in 70 non-redundant nucleic acid binding protein chains solved using X-ray crystallography are analysed. The helical geometry has been characterized as bent, canonical, terminally or completely distorted. The analysis reveals that approximately 70% of the alpha helices possess distortions of any one kind, viz., bend, terminal distortion or complete distortion. Nearly one-third of the total helices possess bends, with a majority of the bending occurring in 5-15 degrees range. In addition, a majority of the bent helices approach the nucleic acid helix in a perpendicular direction. The program RADIL has been useful in characterizing the nucleic acid-induced structural variations in alpha helices, however small they may be.     

Combined nuclear magnetic resonance spectroscopy and molecular dynamics study of growth hormone releasing hexapeptide GHRP‐6 and a cyclic analogue

The Growth Hormone Releasing Hexapeptide, GHRP‐6 was the first of a family of synthetic peptides that enhance the release of the Growth Hormone by the pituitary gland in a dose‐dependent manner. Since its discovery, it has been used as a benchmark and starting point in numerous researches aiming to obtain new drugs. Complete resonance assignment of GHRP‐6 NMR spectra in both open and cyclic forms are reported, showing some differences to random coil chemical shifts. Connectivities observed in the ROESY spectra indicate spatial proximity between the aromatic residues side‐chains in both molecules, as well as between residues DPhe5 and Lys6 sidechains. An ensemble of 10 structures was generated for each one of the molecules, showing RMSD values indicative of nonrandom structures. Molecular Dynamics simulations, both with and without explicit solvent, were carried out for GHRP‐6 and its cyclic analogue. Conformational analysis performed on the trajectories showed a nonrandom structure with a well preserved backbone. The presence of geometrical patterns resembling those typical of π‐π interactions in both peptides, suggest that this kind of interactions may be relevant for the biological activity of GHRP‐6. Same conclusion can be drawn from the spatial proximity of residues DPhe5 and Lys6 sidechains. Copyright © 2012 John Wiley & Sons, Ltd.     

Stabilization centers in various proteins

Interactions among residues together with their interactions with the surrounding medium determine the unique structure of globular proteins. An algorithm was recently developed to locate residues participating in cooperative long-range interactions, called stabilization center residues, that are primarily responsible for preventing the decay of the 3D structure. While our statistical analysis showed that interactions of stabilization center residues hardly influence the formation of the various secondary structure elements, the distribution of the stabilization center residues is rather uneven among the secondary structure elements. Here we analyzed the frequency and distribution of the stabilization center residues and their interacting pairs in secondary structure classes to learn about the effect of secondary structure on the formation and properties of stabilization centers and about the types of interactions responsible for stabilization of proteins of various secondary structure classes. It was found that residues from the same secondary structure tend to interact with each other in the stabilization centers of all classes. It is also suggested that the folding-unfolding equilibrium is governed by different principles for class all-α than for the rest of the classes. Received: 24 April 1998 / Accepted: 17 September 1998 / Published online: 7 December 1998     

圆偏振光在手性聚合物合成与结构调控中的应用

手性是自然界中普遍存在的有趣现象之一,在生命体中手性大分子特有的不对称结构在维持生命过程、新陈代谢和进化等面均起着决定性作用.受此启发,合成具有新型结构的光学活性聚合物,研究其独特的物理化学性质和功能已成为当今高分子领域研究的热点.左旋和右旋的圆偏振光已被广泛应用于氨基酸衍生物的不对称光合成、光分解和去消旋化反应,以及诱导含有偶氮苯或三苯胺等特定功能基团的超分子组装体或无机纳米粒子形成稳定螺旋结构.本文详细地介绍了圆偏振光辐照在手性聚合物合成与螺旋结构调控中的应用,初步揭示了圆偏振光的作用机制以及优势,归纳总结了已取得的研究进展,并对圆偏振光在手性聚合物合成与结构调控中的应用及发展进行了简单的评述和展望.     

Mimicking helical antibacterial peptides with nonpeptidic folding oligomers

Unnatural oligomeric scaffolds designed to adopt defined secondary structures (e.g., helices), while retaining the chemical diversity of amino acid side chains, are of practical value to elaborate functional mimetics of bioactive alpha-polypeptides. Enantiopure N,N'-linked oligoureas as short as seven residues long have been previously shown to fold into a stable helical structure, stabilized by 12- and 14-membered H-bonded rings. We now report that eight-residue oligoureas designed to mimic globally amphiphilic alpha-helical host-defense peptides are effective against both gram-negative and gram-positive bacteria (including methicillin-resistant Staphylococcus aureus [MRSA]) and exhibit selectivity for bacterial versus mammalian cells. Circular dichroism (CD) spectroscopy studies suggest enhanced helical propensity of oligoureas in the presence of phospholipid vesicles. The utility of this new class of nonpeptidic foldamers for biological applications is highlighted by high resistance to proteolytic degradation.     

α N-O turn induced by fluorinated α-aminoxy diamide: synthesis and conformational studies

Two α-aminoxy diamides with fluorinated side chains were synthesized. Their secondary structures characterization was carried out by ¹H NMR, and IR spectrometries as well as X-ray crystallography studies. α N-O turn secondary structures are adopted insusceptibly by side-chain-fluorinated α-aminoxy residues. Thus the fluorinated α-aminoxy diamide can be a potential residue as a biological tracer to be incorporated into aminoxy peptides.     

Aromatic garlands,as new foldamers,to mimic protein secondary structure

Proteins modulate the majority of all biological functions and are composed of highly organized secondary structural elements such as helices, turns, and sheets. Many of these functions are affected by a small number of key structural element, protein–protein interactions. Their mimicry by peptide and non-peptide scaffolds has become a major focus of contemporary research. This paper examines oligomeric system as new foldamers, which either reproduce the local topography of the helix, or project appropriately functionality in a similar manner to residues of an alpha-helix.     

Analysis of molecular structures and mechanisms for toxins derived from venomous animals

As predominant component in the venom of many dangerous animal species, toxins have been thoroughly investigated for drug design or as pharmacologic tools. The present study demonstrated the use of size and hydrophobicity of amino acid residues for the purposes of quantifying the valuable sequence–structure relationship and performing further analysis of interactional mechanisms in secondary structure elements (SSEs) for toxin native conformations. First, we showed that the presence of large and hydrophobic residues varying in availability in the primary sequences correspondingly affects the amount of these residues being used in the SSEs in accordance with linear behavioral patterns from empirical assessments of experimentally derived toxins and non-toxins. Subsequent derivation of prediction rules was established with the aim of analyzing molecular structures and mechanisms by means of 114 residue compositions for venom toxins. The obtained results concerning the linear behavioral patterns demonstrated the nature of the information transfer occurring from the primary to secondary structures. A dual action mechanism was established, taking into account steric and hydrophobic interactions. Finally, a new residue composition prediction method for SSEs of toxins was suggested.     

Constrained Peptides with Target‐Adapted Cross‐Links as Inhibitors of a Pathogenic Protein–Protein Interaction

Bioactive conformations of peptides can be stabilized by macrocyclization, resulting in increased target affinity and activity. Such macrocyclic peptides proved useful as modulators of biological functions, in particular as inhibitors of protein–protein interactions (PPI). However, most peptide‐derived PPI inhibitors involve stabilized α‐helices, leaving a large number of secondary structures unaddressed. Herein, we present a rational approach towards stabilization of an irregular peptide structure, using hydrophobic cross‐links that replace residues crucially involved in target binding. The molecular basis of this interaction was elucidated by X‐ray crystallography and isothermal titration calorimetry. The resulting cross‐linked peptides inhibit the interaction between human adaptor protein 14‐3‐3 and virulence factor exoenzyme S. Taking into consideration that irregular peptide structures participate widely in PPIs, this approach provides access to novel peptide‐derived inhibitors.