Secondary structure inducing potential of beta-amino acids: torsion angle clustering facilitates comparison and analysis of the conformation during MD trajectories

While numerous examples of beta-peptides--exclusively composed of beta-amino acids--have been investigated during the past decade, there are only few reports on the conformational preference of a single beta-amino acid when incorporated into a cyclopeptide. The conformational bias of beta-amino acids on the secondary structure of cyclopeptides has been investigated by NMR spectroscopy in combination with distance geometry (DG) and molecular dynamics (MD) calculations using experimental constraints. The atomic coordinate RMSD criterion usually employed for clustering of conformations after DG and MD calculations does not necessarily group similar peptide conformations, as there is an insufficient correlation between atomic coordinates and torsion angles. To improve on this shortcoming and to eliminate any arbitrary decisions during this process, a torsion angle clustering procedure has been implemented. For the cyclic pentapeptides cyclo-(-Val-beta-Hala-Phe-Leu-Ile-) 1 and cyclo-(-Ser-Pro-Leu-beta-Hasn-Asp-) 3, the beta-amino acid is found in the central position of an extended gamma-turn (pseudo gamma-turn, Psigamma-turn), while the beta-Hpro residue in the cyclic hexapeptide cyclo-(-Ser-beta-Hpro-Leu-Asn-Ile-Asp-) 5 preferentially occupies position i+1 of a pseudo beta-turn (Psibeta-turn). These results further corroborate the hypothesis of beta-amino acids being reliable inducers of secondary structure in cyclic penta- and hexapeptides. They can be employed in the de novo design of biologically active cyclopeptides in pharmaceutical research, since the three-dimensional presentation of pharmacophoric groups in the side chains can be tailored by incorporation of beta-amino acids in strategic sequential positions.     

Spatial structure of myelopeptides: I. Conformational analysis of MP-1, MP-2, and MP-3

Theoretical conformational analysis was used to study the spatial structure and conformational properties of myelopeptides, bone-marrow peptide mediators. The low-energy conformations of three hexapeptides MP-1 (Phe-Leu-Gly-Phe-Pro-Thr), MP-2 (Leu-Val-Val-Tyr-Pro-Trp), and MP-3 (Leu-Val-Cys-Tyr-Pro-Gln) were found, the values of dihedral angles of the backbone and side chains of the amino acid residues constituting these peptides were determined, and the energies of intra- and interresidual interactions were estimated.     

Spatial structure of myelopeptides: I. conformational analysis of MP-1, MP-2, and MP-3

Theoretical conformational analysis was used to study the spatial structure and conformational properties of myelopeptides, bone-marrow peptide mediators. The low-energy conformations of three hexapeptides MP-1 (Phe-Leu-Gly-Phe-Pro-Thr), MP-2 (Leu-Val-Val-Tyr-Pro-Trp), and MP-3 (Leu-Val-Cys-Tyr-Pro-Gln) were found, the values of dihedral angles of the backbone and side chains of the amino acid residues con-stituting these peptides were determined, and the energies of intra- and interresidual interactions were estimated.Translated from Bioorganicheskaya Khimiya, Vol. 31, No. 1, 2005, pp. 31–38.Original Russian Text Copyright © 2005 by Akhmedov, Ismailova, Abbasli, Akhmedov, Godjaev.     

Molecular dynamics simulations of a cyclic-beta-(1-->2) glucan containing an alpha-(1-->6) linkage as a 'molecular alleviator' for the macrocyclic conformational strain

The conformational preferences of a cyclic osmoregulated periplasmic glucan of Ralstonia solanacearum (OPGR), which is composed of 13 glucose units and linked entirely via beta-(1-->2) linkages excluding one alpha-(1-->6) linkage, were characterized by molecular dynamics simulations. Of the three force fields modified for carbohydrates that were applied to select a suitable one for the cyclic glucan, the carbohydrate solution force field (CSFF) was found to most accurately simulate the cyclic molecule. To determine the conformational characteristics of OPGR, we investigated the glycosidic dihedral angle distribution, fluctuation, and the potential energy of the glucan and constructed hypothetical cyclic (CYS13) and linear (LINEAR) glucans. All beta-(1-->2)-glycosidic linkages of OPGR adopted stable conformations, and the dihedral angles fluctuated in this energy region with some flexibility. However, despite the inherent flexibility of the alpha-(1-->6) linkage, the dihedral angles have no transition and are more rigid than that in a linear glucan. CYS13, which consists of only beta-(1-->2) linkages, is somewhat less flexible than other glycans, and one of its linkages adopts a higher energy conformation. In addition, the root-mean-square fluctuation of this linkage is lower than that of other linkages. Furthermore, the potential energy of glucans increases in the order of LINEAR, OPGR, and CYS13. These results provide evidence of the existence of conformational constraints in the cyclic glucan. The alpha-(1-->6)-glycosidic linkage can relieve this constraint more efficiently than the beta-(1-->2) linkage. The conformation of OPGR can reconcile the tendency for individual glycosidic bonds to adopt energetically favorable conformations with the requirement for closure of the macrocyclic ring by losing the inherent flexibility of the alpha-(1-->6)-glycosidic linkage.     

Glaser oxidative coupling on peptides: Stabilization of β-turn structure via a 1,3-butadiyne constraint

The Glaser–Eglinton reaction between either two C or N propargylglycine (Pra or NPra) amino acids, in the presence of copper(II), led to cyclic hexa- and octapeptides constrained by a butadiyne bridge. The on-resin cyclization conditions were analyzed and optimized. The consequences of this type of constraint on the three dimensional structure of these hexapeptides and octapeptides were analyzed in details by NMR and molecular dynamics. We show that stabilized short cyclic peptides could be readily prepared via the Glaser oxidative coupling either with a chiral (Pra), or achiral (NPra) residue. The 1,3-butadiyne cyclization, along with disulfide bridged and lactam cyclized hexapeptides expands the range of constrained peptides that will allow exploring the breathing of amino acids around a β-turn structure.     

Consistent force field calculations on 2,5-diketopiperazine and its 3.6-dimethyl derivatives

The minimum energy conformations are calculated for 2, 5-diketopiperazine (DKP) and its 3,6-dimethyl derivatives (DL-DMDKP and LL-DMDKP), using a consistent force field approach developed previously. The energy function parameters that were not required in earlier calculations on alkanes, amides, mid lactams are fitted to spectral and conformational data on the diketopiperazines. Vibrational assignments are suggested for DKP. Conformational energies are also determined over a range of selected values for ring dihedral angles, and the shape of the potential energy functions is examined over deviations from planarity. DKP and LL-DMDKP are found to have non-planar minimum energy conformations, separated from planar by less than a kcal/mole. DL-DMKP exhibits a nearly flat trough about the planar conformation. Calculations of minimum energies with one dihedral angle coordinate constrainted show a coupling between bond angles and dihedral angles in agreement with recent suggestions of Benedetti.     

Cyclic hexapeptides bearing carboxyl groups. Interaction with metal ions and lipid membrane

Three cyclic hexapeptides bearing carboxyl groups, cyclo(L-Asp-L-Phe-L-Pro)2, cyclo(L-Aad-L-Phe-L-Pro)2 (Aad represents alpha-amino adipic acid residue), and cyclo(D-Asp-D-Phe-L-Pro)2 were synthesized and investigated on conformation, complexation with metal ions, and interaction with lipid membrane. These cyclic hexapeptides were found to take several different conformations, though their reference compounds, cyclo(L-Leu-L-Phe-L-Pro)2 and cyclo[D-Asp(OMe)-D-Phe-L-Pro]2, took a single C2 symmetric conformation. Cyclo(D-Asp-D-Phe-L-Pro)2 formed complexes with Ba2+ and Ca2+, whereas cyclo(L-Asp-L-Phe-L-Pro)2 and cyclo(L-Aad-L-Phe-L-Pro)2 did not. The latter two take amphiphilic structures and were distributed to lipid membrane more easily than cyclo(D-Asp-D-Phe-L-Pro)2. Cyclo(D-Asp-L-Phe-L-Pro)2 binds Ca2+ on the lipid membrane.     

Synthesis and conformation of the cyclic octapeptides cyclo(Phe-Pro)4, cyclo(Leu-Pro)4, and cyclo[Lys(Z)-Pro]4

Cyclic octapeptides, cyclo(X-Pro)₄, where X represents Phe, Leu, or Lys(Z), were synthesized and their conformations investigated. A C₂-symmetric conformer containing two cis peptide bonds was found in all of these cyclic octapeptides. The numbers of available conformations due to the cis–trans isomerization of Pro peptide bonds depended on the nature of the solvent and X residue: they decreased in the following order: cyclo[Lys(Z)-Pro]₄ > cyclo(Leu-Pro)₄ > cyclo(Phe-Pro)₄ in CDCl₃. ¹³C spin-lattice relaxation times (T₁) of these cyclic octapeptides were measured, and the contribution of segmental mobility to T₁ was found to vary with the nature of the X residue.     

Simulation of conformational possibilities of DNA via calculation of nonbonded interactions of complementary dinucleoside phosphate complexes

Using classical potential functions, we carried out potential-energy calculations on the complementary deoxydinucleoside phosphate complexes dApdA:dUpdU, dUpdA:dUpdA, and dApdU:dApdU. All dihedral and bond angles, except those of the nitrogen bases, were varied. The resulting minimum-energy conformations of the complexes are close to DNA A- and B-family conformations, with a typical arrangement of the nitrogen bases. The dihedral and bond angles of one of the molecules forming the complex can thereby differ by several degrees from those of the other molecule. For different base sequences, some dihedral and bond angles may vary over a range of several degrees without appreciably changing the total energy of the complex. Some low-energy conformations of the complexes corresponding to other regions of the conformational space are also found. The biological consequences of possible changes in dihedral and bond angles, occurring on interaction with other molecules, are discussed.     

The effect of conformation on the solution stability of linear vs. cyclic RGD peptides.

The objective of this study was to evaluate the relationship between conformational flexibility and solution stability of a linear RGD peptide (Arg-Gly-Asp-Phe-OH; 1) and a cyclic RGD peptide (cyclo-(1, 6)-Ac-Cys-Arg-Gly-Asp-Phe-Pen-NH2; 2); as a function of pH. Previously, it was found that cyclic peptide 2 was 30-fold more stable than linear peptide 1. Therefore, this study was performed to explain the increase in chemical stability based on the preferred conformation of the peptides. Molecular dynamics simulations and energy minimizations were conducted to evaluate the backbone flexibility of both peptides under simulated pH conditions of 3, 7 and 10 in the presence of water. The reactive sites for degradation for both molecules were also followed during the simulations. The backbone of linear peptide 1 exhibited more flexibility than that of cyclic peptide 2, which was reflected in the rotation about the phi and psi dihedral angles. This was further supported by the low r.m.s. deviations of the backbone atoms for peptide 2 compared with those of peptide 1 that were observed among structures sampled during the molecular dynamics simulations. The presence of a salt bridge between the side chain groups of the Arg and Asp residues was also indicated for the cyclic peptide under simulated conditions of neutral pH. The increase in stability of the cyclic peptide 2 compared with the linear peptide 1, especially at neutral pH, is due to decreased structural flexibility imposed by the ring, as well as salt bridge formation between the side chains of the Arg and Asp residues in cyclic peptide 2. This rigidity would prevent the Asp side chain carboxylic acid from orienting itself in the appropriate position for attack on the peptide backbone.     

Local propensities and statistical potentials of backbone dihedral angles in proteins

The following three issues concerning the backbone dihedral angles of protein structures are presented. (1) How do the dihedral angles of the 20 amino acids depend on the identity and conformation of their nearest residues? (2) To what extent are the native dihedral angles determined by local (dihedral) potentials? (3) How to build a knowledge-based potential for a residue's dihedral angles, considering the identity and conformation of its nearest residues? We find that the dihedral angle distribution for a residue can significantly depend on the identity and conformation of its adjacent residues. These correlations are in sharp contrast to the Flory isolated-pair hypothesis. Statistical potentials are built for all combinations of residue triplets and depend on the dihedral angles between consecutive residues. First, a low-resolution potential is obtained, which only differentiates between the main populated basins in the dihedral angle density plots. Minimization of the dihedral potential for 125 test proteins reveals that most native alpha-helical residues (89%) and a large fraction of native beta-sheet residues (47%) adopt conformations close to their native one. For native loop residues, the percentage is 48%. It is also found that this fraction is higher for residues away from the ends of alpha or beta secondary structure elements. In addition, a higher resolution potential is built as a function of dihedral angles by a smoothing procedure and continuous functions interpolations. Monte Carlo energy minimization with this potential results in a lower fraction for native beta-sheet residues. Nevertheless, because of the higher flexibility and entropy of beta structures, they could be preferred under the influence of non-local interactions. In general, most alpha-helices and many beta-sheets are strongly determined by the local potential, while the conformations in loops and near the end of beta-sheets are more influenced by non-local interactions.     

Ab initio computational study of beta-cellobiose conformers using B3LYP/6-311++G**

The molecular structure of 27 conformers of beta-cellobiose were studied in vacuo through gradient geometry optimization using B3LYP density functionals and the 6-311++G** basis set. The conformationally dependent geometry changes and energies were explored as well as the hydrogen-bonding network. The lowest electronic energy structures found were not those suggested from available crystallographic and NMR solution data, where the glycosidic dihedral angles fall in the region (phi, psi) approximately (40 degrees, -20 degrees ). Rather, 'flipped' conformations in which the dihedral angles are in the range (phi, psi) approximately (180 degrees, 0 degrees ) are energetically more stable by approximately 2.5 kcal/mol over the 'experimentally accepted' structure. Further, when the vibrational free energy, deltaG, obtained from the calculated frequencies, is compared throughout the series, structures with (phi, psi) in the experimentally observed range still have higher free energy ( approximately 2.0 kcal/mol) than 'flipped' forms. The range of bridging dihedral angles of the 'normal' conformers, resulting from the variance in the phi dihedral is larger than that found in the 'flipped' forms. Due to this large flat energy surface for the normal conformations, we surmise that the summation of populations of these conformations will favor the 'normal' conformations, although evidence suggests that polar solvent effects may play the dominant role in providing stability for the 'normal' forms. Even though some empirical studies previously found the 'flipped' conformations to be lowest in energy, these studies have been generally discredited because they were in disagreement with experimental results. Most of the DFT/ab initio conformations reported here have not been reported previously in the ab initio literature, in part because the use of less rigorous theoretical methods, i.e. smaller basis sets, have given results in general agreement with experimental data, that is, they energetically favored the 'normal' forms. These are the first DFT/ab initio calculations at this level of theory, apparently because of the length and difficulty of carrying out optimizations at these high levels.     

Analogues of Somatostatin Bind Selectively to Brain Somatostatin Receptor Subtypes

Somatostatin (SRIF) is a neurotransmitter that produces its multiple effects in the CNS through interactions with membrane-bound receptors. Subtypes of SRIF receptors are found in the CNS that are distinguished by their sensitivities to the cyclic hexapeptide MK-678, such that SRIF1 receptors are sensitive to MK-678 and SRIF2 receptors are insensitive to MK-678. In the present study, we further examined the selectivities of a series of structurally diverse SRIF analogues for SRIF receptor subtypes. SRIF receptors were labeled by 125I-Tyr11-SRIF, which has indistinguishable affinities for SRIF receptor subtypes. The inhibition by MK-678 was incomplete, indicating this peptide is highly selective for a subtype of SRIF receptor that we have termed the SRIF1 receptor. The binding of 125I-MK-678 to SRIF1 receptors was monophasically inhibited by SRIF, the octapeptides (such as SMS-201-995), and the hexapeptides (such as MK-678), consistent with the highly selective labeling of a subtype of SRIF receptor. In contrast, the smaller CGP-23996-like analogues did not inhibit 125I-MK-678 binding to SRIF1 receptors. The binding of 125I-CGP-23996 to SRIF receptors was inhibited by SRIF and the octapeptides with Hill coefficients of less than 1, indicating that 125I-CGP-23996 labels multiple SRIF receptor subtypes. The hexapeptides and CGP-23996-like compounds produced only partial inhibitions of 125I-CGP-23996 binding, which were additive, indicating selective interactions of these compounds with the different receptor subpopulations labeled by 125I-CGP-23996. 125I-Tyr11-SRIF binding and 125I-CGP-23996 binding to SRIF receptors were likewise only partially affected by 100 microM guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S), a concentration that completely abolishes specific 125I-MK-678 binding to SRIF1 receptors. The component of 125I-CGP-23996 labeling that was sensitive to GTP gamma S was also MK-678 sensitive. Thus, two subpopulations of SRIF receptors exist in the CNS. The SRIF1 receptor is sensitive to cyclic hexapeptides such as MK-678 and to GTP gamma S but insensitive to smaller CGP-23996-like compounds. The SRIF2 receptor is sensitive to the CGP-23996-like compounds and can be selectively labeled by 125I-CGP-23996 in the presence of high concentrations of the hexapeptides or GTP gamma S because, unlike the SRIF1 receptor, the SRIF2 receptor is insensitive to these agents. The SRIF receptor subtype-selective peptide analogues will be useful in the future characterization of the functions mediated by SRIF receptor subtypes in the CNS.     

Ca2+ transport through lipid membrane by diastereomer cyclic octapeptides

Effects of the nature and orientation of a side chain in cyclic octapeptides on Ca2+ transport were examined by using cyclo[L-Lys(Z)-Sar-L-Leu-Sar]2 (C8-L), cyclo[L-Lys(Z)-Sar]4 (C8KS), and their diastereomer cyclic octapeptides, cyclo[L-Lys(Z)-Sar-D-Leu-Sar]2 (C8-D) and cyclo[L-Lys(Z)-Sar-D-Lys(Z)-Sar]2 (C8Kk). All these cyclic octapeptides were found to take a single conformation in CDCl3, and the conformation was C2-symmetric for C8-L and C8-D, and C4-symmetric for C8KS and C8Kk. They formed a complex with Ca2+. Upon complexation, C8KS accompanied isomerization of peptide bonds, but C8-D retained the arrangement of peptide bonds. The amount of Ca2+ extracted from an aqueous solution to a chloroform solution by all L cyclic octapeptide C8-L or C8KS was about twice that of Na+, but 6-8-fold smaller than that by C8-D or C8Kk including D units. These cyclic octapeptides were capable of transporting Ca2+ through a lipid membrane above the phase transition temperature, and the transport rate decreased in the order of C8Kk-C8KS greater than C8-D greater than C8-L.     

β-Branched Amino Acids Stabilize Specific Conformations of Cyclic Hexapeptides

Cyclic peptides (CPs) are a promising class of molecules for drug development, particularly as inhibitors of protein-protein interactions. Predicting low-energy structures and global structural ensembles of individual CPs is critical for the design of bioactive molecules, but these are challenging to predict and difficult to verify experimentally. In our previous work, we used explicit-solvent molecular dynamics simulations with enhanced sampling methods to predict the global structural ensembles of cyclic hexapeptides containing different permutations of glycine, alanine, and valine. One peptide, cyclo-(VVGGVG) or P7, was predicted to be unusually well structured. In this work, we synthesized P7, along with a less well-structured control peptide, cyclo-(VVGVGG) or P6, and characterized their global structural ensembles in water using NMR spectroscopy. The NMR data revealed a structural ensemble similar to the prediction for P7 and showed that P6 was indeed much less well-structured than P7. We then simulated and experimentally characterized the global structural ensembles of several P7 analogs and discovered that β-branching at one critical position within P7 is important for overall structural stability. The simulations allowed deconvolution of thermodynamic factors that underlie this structural stabilization. Overall, the excellent correlation between simulation and experimental data indicates that our simulation platform will be a promising approach for designing well-structured CPs and also for understanding the complex interactions that control the conformations of constrained peptides and other macrocycles.     

Analysis of the relationship between side-chain conformation and secondary structure in globular proteins

The relationship between the preferred side-chain dihedral angles and the secondary structure of a residue was examined. The structures of 61 proteins solved to a resolution of 2.0 A (1 A = 0.1 nm) or better were analysed using a relational database to store the information. The strongest feature observed was that the chi 1 distribution for most side-chains in an alpha-helix showed an absence of the g- conformation and a shift towards the t conformation when compared to the non-alpha/beta structures. The exceptions to this tendency were for short polar side-chains that form hydrogen bonds with the main-chain which prefer g+. Shifts in the chi 1 preferences for residues in the beta-sheet were observed. Other side-chain dihedral angles (chi 2, chi 3, chi 4) were found to be influenced by the main-chain. This paper presents more accurate distributions for the side-chain dihedral angles which were obtained from the increased number of proteins determined to high resolution. The means and standard deviations for chi 1 and chi 2 angles are presented for all residues according to the secondary structure of the main-chain. The means and standard deviations are given for the most popular conformations for side-chains in which chi 3 and chi 4 rotations affect the position of C atoms.     

DFT conformation and energies of amylose fragments at atomic resolution. Part 2: ‘band-flip’ and ‘kink’ forms of α-maltotetraose

In Part 2 of this series of DFT optimization studies of α-maltotetraose, we present results at the B3LYP/6-311++G∗∗ level of theory for conformations denoted ‘band-flips’ and ‘kinks’. Recent experimental X-ray studies have found examples of amylose fragments with conformations distorted from the usual syn forms, and it was of interest to examine these novel structural motifs by the same high-level DFT methods used in Part 1. As in Part 1, we have examined numerous hydroxymethyl rotamers (gg, gt, and tg) at different locations in the residue sequence, and include the two hydroxyl rotamers, the clockwise ‘c’ and counterclockwise ‘r’ forms. A total of fifty conformations were calculated and energy differences were found to attempt to identify those sources of electronic energy that dictate stressed amylose conformations. Most stressed conformations were found to have relative energies considerably greater (i.e., ∼4 to 12 kcal/mol) than the lowest energy syn forms. Relative energy differences between ‘c’ and ‘r’ forms are somewhat mixed with some stressed conformations being ‘c’ favored and some ‘r’ favored, with the lowest energy ‘kink’ form being an all-gg-r conformation with the ‘kink’ in the bc glycosidic dihedral angles. Comparison of our calculated structures with experimental results shows very close correspondence in dihedral angles.     

Spatial structure of isoleucine pentapeptides Glu-Phe-Leu-Arg-Ile-NH2 and Pro-Phe-Tyr-Arg-Ile-NH2

The spatial structure of two cardioactive isoleucine pentapeptides Glu-Phe-Leu-Arg-Ile-NH2 (I) and Pro-Phe-Tyr-Arg-Ile-NH2 (II) have been investigated using the theoretical conformational analysis. The low-energy conformations of these molecules were found, the values of dihedral angles of the backbone and side chains of the amino acid residues constituting these peptides were determined, and the energies of intra- and interresidual interactions were estimated. It was revealed that the spatial structure of molecule I can exist as five and that of molecule II as seven stable backbone forms.     

Solvent accessibility studies of β-bends and application to cyclic hexapeptides

The solvent-accessible surface areas (ASAs), of the atoms in tripeptides around the minimum-energy conformations of the β-bend types I, I′, II, and II′ have been computed as a first step in the systematic solvent accessibiity study of secondary structures. The side chains chosen at the two middle positions of the bend are L -Ala, D -Ala, and Gly. The ASAs of the hydrogen atoms are reported here and are found useful in determining the type of β-bends in six examples of cyclic hexapeptides whose crystal structures are known. Comparison with observation showed that all the β-bends in these cyclic hexapeptides were correctly identified by the present method. This points to a possible use of the method in identifying β-bend types in solution.     

Chain reversals in model peptides: studies of cystine-containing cyclic peptides. II. Effects of valyl residues and possible i-to-(i + 3) attractive ionic interactions on cyclization of [Cys1], [Cys6] hexapeptides

The synthesis of four N-acetyl N'-methylamide cystine-containing hexapeptides, CVPGVC, CGVVGC, CKPGEC, and CEPGKC, is described. These were used in disulfide-exchange reactions with the peptide CVPGGC as the formal oxidant. The relative propensities for peptide cyclization were thus deduced, and the tendency toward the formation of chain-reversal conformations was established quantitatively. An additional peptide, CVVVVC, was prepared but was never obtained as the cyclic monomer, demonstrating that the formation of chain-reversals in this peptide was of very low probability. Incorporation of pairs of valyl residues decreased the ease of cyclization, but it appeared that conformational flexibility in the cystine-containing hexapeptides may have compensated for substitutions which would have been expected to hinder the adoption of certain beta-turn conformations. The peptides containing ionic residues were cyclized more readily than expected, and this process was relatively insensitive to salt concentration. This observation is discussed with regard to the stabilization of beta-turns by i-to-(i + 3) ionic interactions in peptides and proteins. A method for blocking thiols was introduced as an improvement in the analysis of the equilibrium mixtures.