Conformational studies of retro–inverso peptides: The crystal and molecular structure of the hydantoin from H-Ala-G-Ala-mGly-OBzl

The crystal structure of the hydantoin 1-[(S)-1′–aminoethylmalonyl benzyl ester]-(S)-4-methylimidazolidin-2.5-dione (1) derived from the peptide H-Ala-gAla-mGly-OBzl, Having the retro-inverso modification of the Ala-Gly bond, has been determined by x-ray diffraction analysis. The crystals are orthorhombic, space group P2₁2₁2₁ with a = 6.539. b = 14.721, c = 17.101 Å, z = 4. The structure was solved by direct methods and refined with anisotropic thermal factors to a final R value of 0.067 for the 947 observed reflections. Reversal of the Ala-Gly amide bond perturbs the folding tendency of the backbone shown by the parent peptide t-BuCO-Ala-Gly-NHiPr. The gem-diamino residue, gAla, and the malonyl moieties are found in the helical and the extended conformations, respectively. Intramolecular hydrogen bonding is not observed. The molecules in the crystal are held together by the formation of two intermolecular hydrogen bonds of the N? H?O?C type with N?O distances of 2.86 and 3.17 Å respectively. 1995 John Wiley&Sons. Inc. © 1995 John Wiley & Sons, Inc.     

Conformational characteristics of unstructured peptides: alpha-synuclein

We have performed replica-exchange molecular dynamics simulations on 41 residue peptides containing NAC region of alpha-synuclein in various force fields and solvent conditions. Alpha-synuclein is known to be the major cause of Parkinson's disease by amyloid-like aggregation, and one of the natively unfolded proteins. To investigate conformational characteristics of intrinsically unstructured peptides, we carried out structural analysis by introducing 'representative structure' for ensemble of structures occurring during the overall trajectory. Representative structures may be defined by using either coordinate averaging or distance averaging. When applied to the natively folded proteins such as villin headpiece, structural analysis based on representative structure was found to yield consistent results with those obtained from conventional analysis. Individual conformations obtained from the simulations of NAC peptide for various conditions show flexible structures close to random coil. Secondary structure contents and free energy surfaces showed dependency on solvent conditions, which may be interpreted as another manifestation of structural diversity. It is found that representative structures can provide useful information about structural characteristics of intrinsically unstructured proteins.     

Conformational states and folding pathways of peptides revealed by principal-independent component analyses

Principal component analysis is a powerful method for projecting multidimensional conformational space of peptides or proteins onto lower dimensional subspaces in which the main conformations are present, making it easier to reveal the structures of molecules from e.g. molecular dynamics simulation trajectories. However, the identification of all conformational states is still difficult if the subspaces consist of more than two dimensions. This is mainly due to the fact that the principal components are not independent with each other, and states in the subspaces cannot be visualized. In this work, we propose a simple and fast scheme that allows one to obtain all conformational states in the subspaces. The basic idea is that instead of directly identifying the states in the subspace spanned by principal components, we first transform this subspace into another subspace formed by components that are independent of one other. These independent components are obtained from the principal components by employing the independent component analysis method. Because of independence between components, all states in this new subspace are defined as all possible combinations of the states obtained from each single independent component. This makes the conformational analysis much simpler. We test the performance of the method by analyzing the conformations of the glycine tripeptide and the alanine hexapeptide. The analyses show that our method is simple and quickly reveal all conformational states in the subspaces. The folding pathways between the identified states of the alanine hexapeptide are analyzed and discussed in some detail.     

The crystal structure of Afc-containing peptides

A systematic structural analysis of Afc (9-amino-fluorene-9-carboxylic acid) containing peptides is here reported. The crystal structures of four fully protected tripeptides containing the Afc residue in position 2: Z-X(1)-Afc(2)-Y(3)-OMe (peptide a: X = Y = Gly; peptide b: X = Aib, C(alpha, alpha)-dimethylglycine, Y = Gly; peptide c: X = Gly, Y = Aib; peptide d: X = Y = Aib) have been solved by x-ray crystallography. All the results suggest that the Afc residue has a high propensity to assume an extended conformation. In fact, the Afc residue adopts an extended conformation in three peptides examined in this paper (peptides a-c). In contrast, Afc was found in a folded conformation, in the 3(10)-helical region, only in the peptide d, in which it is both preceded and followed by the strong helix promoting Aib.     

Conformational difference between diastereomers of Dnp-Val-Aib-Gly-Leu-pNA studied by x-ray crystal analyses

In order to investigate the Conformational change of the α-aminoisobutyric acid (Aib) containing peptide by the D /L replacement of an amino acid residue, single crystals of two diastereomers, Dnp-L -Val-Aib-Gly-L -Leu-pNA (L -L isomer) and Dnp-D -Val-Aib-Gly-L -Leu-pNA (D -L isomer), were prepared from aqueous methanol solutions as CH₃OH and CH₃OH · H₂O solvates, respectively, and were analyzed by the x-ray diffraction method. Molecular conformation of L -L isomer adopts consecutive two different types of β-turns, a type II′ β-turn bent at Aib-Gly, and a type III β-turn bent at Gly-Leu, stabilized by two intramolecular (Leu) NH …? O?C (Val) and (pNA) NH …? O?C(Aib) hydrogen bonds. In contrast, these two intramolecular hydrogen bonds lead the D -L isomer to a distorted 3₁₀-helix conformation consisting of consecutive two type-III β-turn of Aib-Gly-Leu sequence. The most significant structural difference between these diastereomers is the mutual orientation between the Dnp and pNA chromophores. While the extensive stacking of both the chromophores is intramolecularly formed for the folded conformation of L -L isomer, they are oriented toward an opposite direction in the open conformation of D -L isomer and are intermolecularly stacked with each other. The large separation between these diastereomers observed in the chromatography is discussed in the relation with their Conformational differences. © 1993 John Wiley & Sons, Inc.     

Conformational and structural characteristics of peptides binding to HLA-DR molecules.

A fundamental characteristic of MHC class I and class II proteins is their unusual capacity to form stable complexes with a wide spectrum of peptide ligands. In this study, sets of peptide analogues containing long chain-biotinylated lysine individually substituted for each amino acid in the sequence have been used to explore the structural requirements for the formation of peptide-MHC class II protein complexes. Based on the ability of the analogs to bind both the MHC protein and fluorescent streptavidin, receptor contact residues were identified and from their spacing the conformation of the bound peptides could be inferred. Six separate peptides were studied; three defined by HLA-DR1Dw1-restricted T cells, and three identified by T cells restricted through alleles other than HLA-DR1Dw1. The similar patterns of fluorescent signals observed when the former three peptides were studied indicated that they shared conformational features when bound to HLA-DR1Dw1. In contrast when the latter three peptides were examined, the data indicated that they shared some but not all of the conformational features characteristic of the peptides known to elicit HLA-DR1Dw1-restricted T cells. When the peptide sequences were aligned based on the critical contact residues, two positions of structural homology were apparent. In each sequence, an amino acid with a bulky hydrophobic side chain could be identified separated by four residues from a small amino acid. These minimal structural requirements were consistent with recent experiments demonstrating that only a small number of side chains in the peptide were necessary for binding to the MHC protein.     

Conformational flexibility revealed by the crystal structure of a crenarchaeal RadA

Homologous recombinational repair is an essential mechanism for repair of double-strand breaks in DNA. Recombinases of the RecA-fold family play a crucial role in this process, forming filaments that utilize ATP to mediate their interactions with single- and double-stranded DNA. The recombinase molecules present in the archaea (RadA) and eukaryota (Rad51) are more closely related to each other than to their bacterial counterpart (RecA) and, as a result, RadA makes a suitable model for the eukaryotic system. The crystal structure of Sulfolobus solfataricus RadA has been solved to a resolution of 3.2 Å in the absence of nucleotide analogues or DNA, revealing a narrow filamentous assembly with three molecules per helical turn. As observed in other RecA-family recombinases, each RadA molecule in the filament is linked to its neighbour via interactions of a short β-strand with the neighbouring ATPase domain. However, despite apparent flexibility between domains, comparison with other structures indicates conservation of a number of key interactions that introduce rigidity to the system, allowing allosteric control of the filament by interaction with ATP. Additional analysis reveals that the interaction specificity of the five human Rad51 paralogues can be predicted using a simple model based on the RadA structure.     

Prothrombin structure: unanticipated features and opportunities

The structure of prothrombin has eluded investigators for decades but recent efforts have succeeded in revealing the architecture of this important clotting factor. Unanticipated features have emerged outlining the significant flexibility of the zymogen due to linker regions connecting the γ carboxyglutamic domain, kringles and protease domain. A new, structure-based framework helps in defining a molecular mechanism of prothrombin activation, rationalizes the severe bleeding phenotypes of several naturally occurring mutations and identifies targets for drug design.     

Observation of glycine zipper and unanticipated occurrence of ambidextrous helices in the crystal structure of a chiral undecapeptide

Background  

The de novo design of peptides and proteins has recently surfaced as an approach for investigating protein structure and function. This approach vitally tests our knowledge of protein folding and function, while also laying the groundwork for the fabrication of proteins with properties not precedented in nature. The success of these studies relies heavily on the ability to design relatively short peptides that can espouse stable secondary structures. To this end, substitution with α, β-dehydroamino acids, especially α, β-dehydrophenylalanine (ΔPhe) comes in use for spawning well-defined structural motifs. Introduction of ΔPhe induces β-bends in small and 3₁₀-helices in longer peptide sequences.     

Conformational changes of cholesteryl palmitoleate in the crystal structure at low temperature

At 123 K, crystals of cholesteryl cis-9-hexadecenoate (cholesteryl palmitoleate, C45H74O2) are monoclinic, space group P2(1) with cell dimensions a = 12.917(7), b = 8.910(5), c = 34.04(1) A, beta = 94.95(7) degrees [lambda(CuK alpha) = 1.5424 A] having two independent molecules (A and B) per unit cell. The crystal structure has been determined from 6178 reflections with sin theta/lambda less than or equal to 0.56 A-1, of which 3406 gave [F] greater than 3 sigma. Structure refinement by alternating cycles of Fourier syntheses and block diagonal least squares gave R = 0.24 for all reflections, R = 0.13 for reflections [F] greater than 3 sigma. At 123 K, the crystal structure consists of closely packed layers very similar to those at 295 K. However, there are major conformational differences in the layer interface region, which affect the ester chain of molecule B and the C(17) tail of molecule A. Although the electron density is diffuse in this region, the B-chain, which is bent, appears to be ordered at 123 K and has a different conformation from the disordered B-chains at 295 K. The change in the A-tail, which is twisted at 123 K and extended at 295 K, is very similar to that which occurs in two of the molecules when anhydrous cholesterol undergoes phase transition. Measurements of the unit cell dimensions at twelve temperatures (295 K to 123 K) indicate that the major changes in the crystal structure of cholesteryl palmitoleate occur in a 10 K range near 173 K.     

Solid-state conformations of aminosuccinyl peptides: crystal structure of tert-butyloxycarbonyl-L-leucyl-L-aminosuccinyl-L-phenylalaninami de

The protected tripeptide tert-butyloxycarbonyl-L-leucyl-L-aminosuccinyl-L-phenylalaninamide crystallizes in the orthorhombic space group P2(1)2(1)2(1), with a = 6.214(3), b = 12.832(3), c = 33.094(4) A, Z = 4. The structure was solved by direct methods using MULTAN 80 and refined to an R value of 0.055 for 1458 reflections. The bond lengths and angles are in good agreement with the standard values. The peptide backbone adopts a type II' beta-bend conformation with a weak intramolecular hydrogen bond between the CO group of the leucyl residue and the C-terminal NH2 group. In agreement with previous studies, this structure confirms the high propensity of aminosuccinyl peptides to adopt a type II' beta-bend conformation. The role of this conformation in relation to the deamidation process in proteins is also discussed.     

Conformational analyses of thiirane-based gelatinase inhibitors

SB-3CT is a thiirane-containing inhibitor of the gelatinase class of matrix metalloprotease enzymes. In support of the mechanistic study of this inhibition, the conformational analyses of SB-3CT (and of two methyl-substituted derivatives) were undertaken using X-ray crystallography and molecular dynamics simulation.     

Conformational behavior of ionic self-complementary peptides

Several de novo designed ionic peptides that are able to undergo conformational change under the influence of temperature and pH were studied. These peptides have two distinct surfaces with regular repeats of alternating hydrophilic and hydrophobic side chains. This permits extensive ionic and hydrophobic interactions resulting in the formation of stable beta-sheet assemblies. The other defining characteristic of this type of peptide is a cluster of negatively charged aspartic or glutamic acid residues located toward the N-terminus and positively charged arginine or lysine residues located toward the C-terminus. This arrangement of charge balances the alpha-helical dipole moment (C --> N), resulting in a strong tendency to form stable alpha-helices as well. Therefore, these peptides can form both stable alpha-helices and beta-sheets. They are also able to undergo abrupt structural transformations between these structures induced by temperature and pH changes. The amino acid sequence of these peptides permits both stable beta-sheet and alpha-helix formation, resulting in a balance between these two forms as governed by the environment. Some segments in proteins may also undergo conformational changes in response to environmental changes. Analyzing the plasticity and dynamics of this type of peptide may provide insight into amyloid formation. Since these peptides have dynamic secondary structure, they will serve to refine our general understanding of protein structure.     

Conformational studies of peptides. The crystal structures of N-acetyl-L-prolinamide and N-acetyl-(S)-thiazolidine-4-carboxamide

The crystal structure of N′-acetyl-L -prolinamide and its isomorphous alalog N′-acetyl-(S)-thiazolidine-4-carboxamide was determined using highly accurate parameters obtained by room- and low-temperature data-collecting systems. Both crystals are orthorhombic, space group P2₁2₁2₁, with four molecules per unit cell held together by a hydrogen-bond system that extends in all directions. Both molecules exhibit the following conformational features: the acetyl group is in the trans configuration in respect to the tertiary amide. The primary amide is almost at right angles with respect to the mean plane of the ring and the –NH₂ group is over the ring. The pyrrolidine and thiazolidine rings were found to be rather flexible with C^β puckering in the former and sulfur in the latter.     

Conformational analysis corresponding to intra-chain disulfide bridged peptides in proteins of known three-dimensional structure

We have carried out a systematic analysis in order to evaluate whether Intra-Chain Disulfide Bridged Peptides (ICDBPs) observed in proteins of known three-dimensional structure adopt structurally similar conformations as they may correspond to structural/functional motifs. 406 representative ICDBPs comprising between 3 to 17 amino acid residues could be classified according to peptide sequence length and main-chain secondary structure conformation into 146 classes. ICDBPs comprising 6 amino acid residues are maximally represented in the Protein Data Bank. They also represent the maximum number of main-chain secondary structure conformational classes. Individual ICDBPs in each class represent different protein superfamilies and correspond to different amino acid sequences. We identified 145 ICDBP pairs that had not less-than 0.5 A root mean square deviation value corresponding to their equivalent peptide backbone atoms. We believe these ICDBPs represent structural motifs and possible candidates in order to further explore their structure/function role in the corresponding proteins. The common conformational classes observed for ICDBPs defined according to the main-chain secondary structure conformations; H (helix), B (residue in a isolated beta bridge), C (coil), E (extended beta strand), G (3(10) helix), I (pi helix), S (bend), T (hydrogen-bonded turn) were; "CHHH", "CTTC", "CSSS" and "CSSC" (for ICDBP length 4), "CSSCC" (length 5), "EETTEE", "CCSSCC", "CCSSSC" (length 6), "EETTTEE" (length 7), "EETTTTEE" (length 8), "EEEETTEEEE" (length 10), "EEEETTTEEEE" (length 11) and "EEEETTTTEEEE" (length 12).     

Conformational characteristics and correlations in crystal structures of nucleic acid oligonucleotides: evidence for sub-states

Sugar phosphate backbone conformations are a structural element inextricably involved in a complete understanding of specific recognition nucleic acid ligand interactions, from early stage discrimination of the correct target to complexation per se, including any structural adaptation on binding. The collective results of high resolution DNA, RNA and protein/DNA crystal structures provide an opportunity for an improved and enhanced statistical analysis of standard and unusual sugar-phosphate backbone conformations together with corresponding dinucleotide sequence effects as a basis for further exploration of conformational effects on binding. In this study, we have analyzed the conformations of all relevant crystal structures in the nucleic acids data base, determined the frequency distribution of all possible epsilon, zeta, alpha, beta and gamma backbone angle arrangements within four nucleic acid categories (A-RNA and A-DNA, free and bound B-DNA) and explored the relationships between backbone angles, sugar puckers and selected helical parameters. The trends in the correlations are found to be similar regardless of the nucleic acid category. It is interesting that specific structural effects exhibited by the different unusual backbone sub-states are in some cases contravariant. Certain alpha/gamma changes are accompanied by C3' endo (north) sugars, small twist angles and positive values of base pair roll, and favor a displacement of nucleotide bases towards the minor groove compared to that of canonical B form structures. Unusual epsilon/zeta combinations occur with C2' (south) sugars, high twist angles, negative values of base pair roll, and base displacements towards the major groove. Furthermore, any unusual backbone correlates with a reduced dispersion of equilibrium structural parameters of the whole double helix, as evidenced by the reduced standard deviations of almost all conformational parameters. Finally, a strong sequence effect is displayed in the free oligomers, but reduced somewhat in the ligand bound forms. The most variable steps are GpA and CpA, and, to a lesser extent, their partners TpC and TpG. The results provide a basis for considering if the variable and non-variable steps within a biological active sequence precisely determine morphological structural features as the curvature direction, the groove depth, and the accessibility of base pair for non covalent associations.     

Conformational analysis of two cyclic disulfide peptides

Complete nmr and CD studies of two cyclic tetrapeptides with disulfide bonds, Ac-L-Pen-L-Pro-D-Val-L-Cys-NH2 (1) and Ac-L-Cys-L-Pro-D-Val-L-Cys-NH2 (2) bonds have been carried out in different solvents to investigate the formation and stabilization of beta-turn structures and to determine the stereochemistry of the disulfide linkage. Both peptides have three-dimensional structures with a type II beta-turn, as derived from quantitative nuclear Overhauser effect data. The combined use of CD and nmr indicates that the dihedral angle of the disulfide bridge is different in the two peptides, although the chirality is maintained.     

Conformational analysis of cyclic peptides in solution

The strategy and tactics of conformational analysis of cyclic peptides in solution is demonstrated by the example of cyclo(-D-Pro-Phe-Thr-Phe-Trp-Phe-). Spin-locked experiments like rotating frame nuclear Overhauser enhancement spectroscopy (ROESY), ROTO, and TOCSY are successfully applied to assign all proton signals and to obtain distance information. A crude conformational model was built using the nmr data. This starting model was refined by restrained molecular dynamics (MD) calculations using ROE derived distances and fixed bond angles as determined from homo- and heteronuclear coupling constants. To mimic the solvent and to reduce artifacts in an in vacuo calculation the charges of the solvent-exposed NH protons were gradually reduced according to the temperature gradients. The thus obtained "conformation" (mean of a 40 ps MD trajectory) shows very close similarity to x-ray structures in an orthorhombic and in two monoclinic crystal modifications of the same compound. The main difference is the breaking of an intermolecular hydrogen bond of the threonine hydroxyl group on dissolution of the crystal and forming an intramolecular hydrogen bond in solution.