Distribution Of Side-chain Rotamers Research Articles

Modeling the role of charged residues in thermophilic proteins by rotamer and dynamic cross correlation analysis.

Discerning the determinants of protein thermostability is very important both from the theoretical and applied perspective. Different lines of evidence seem to indicate that a dynamical network of salt bridges/charged residues plays a fundamental role in the thermostability of enzymes. In this work, we applied measures of dynamic variance, like the Gini coefficients, Kullback-Leibler (KL) divergence and dynamic cross correlation (DCC) coefficients to compare the behavior of 3 pairs of homologous proteins from the thermophilic bacterium Thermus thermophilus and mesophilic Escherichia coli. Molecular dynamic (MD) simulations of these proteins were performed at 303K and 363K. In the characterization of their side chain rotamer distributions, the corresponding Gini coefficients and KL-divergence both revealed significant correlations with temperature. Similarly, a DCC analysis revealed a higher trend to de-correlate the movement of charged residues at higher temperatures in the thermophilic proteins, when compared with their mesophilic homologues. These results highlight the importance of dynamic electrostatic network interactions for the thermostability of enzymes.

The interplay between transient α-helix formation and side chain rotamer distributions in disordered proteins probed by methyl chemical shifts

The peptide backbones of disordered proteins are routinely characterized by NMR with respect to transient structure and dynamics. Little experimental information is, however, available about the side chain conformations and how structure in the backbone affects the side chains. Methyl chemical shifts can in principle report the conformations of aliphatic side chains in disordered proteins and in order to examine this two model systems were chosen: the acid denatured state of acyl-CoA binding protein (ACBP) and the intrinsically disordered activation domain of the activator for thyroid hormone and retinoid receptors (ACTR). We find that small differences in the methyl carbon chemical shifts due to the γ-gauche effect may provide information about the side chain rotamer distributions. However, the effects of neighboring residues on the methyl group chemical shifts obscure the direct observation of γ-gauche effect. To overcome this, we reference the chemical shifts to those in a more disordered state resulting in residue specific random coil chemical shifts. The (13)C secondary chemical shifts of the methyl groups of valine, leucine, and isoleucine show sequence specific effects, which allow a quantitative analysis of the ensemble of χ(2)-angles of especially leucine residues in disordered proteins. The changes in the rotamer distributions upon denaturation correlate to the changes upon helix induction by the co-solvent trifluoroethanol, suggesting that the side chain conformers are directly or indirectly related to formation of transient α-helices.

Influence of Side Chain Conformations on Local Conformational Features of Amino Acids and Implication for Force Field Development

Statistical analysis of coil regions in protein structures has been used to obtain the local backbone phi, psi preferences of amino acids, which agree well with the NMR experiments of unfolded peptides and proteins. We analyzed the conformational features of amino acid residues in a restricted coil library of 4220 high-resolution protein crystal structures. In addition to Gly, Ala, and Pro, the phi, psi distribution (Ramachandran plot) of each amino acid is analyzed with respect to three side chain conformers: g+ (chi(1) approximately -60 degrees), g- (chi(1) approximately 60 degrees), and t (chi(1) approximately 180 degrees). The statistical study indicates that the effect of side chain conformations on phi, psi distributions is even greater than the effect of amino acid types. On the basis of the chi(1), phi, psi conformational preferences, the amino acids in addition to Gly, Pro, and Ala can be divided into five types: (1) ordinary amino acids, (2) Ser, (3) Asp and Asn, (4) Val and Ile, and (5) Thr, each with distinguished chi(1) rotamers. The alpha-helix, beta-sheet, and type-I beta-turn preferences of the different rotamers of various amino acid types can be captured by their intrinsic phi, psi preferences from our coil library. Molecular dynamics simulations of dipeptide Ac-X-NHMe and tetrapeptide Ac-A-X-A-NHMe models give nearly the same side chain rotamer distributions. However, for many amino acids, both OPLS-AA/L and AMBER-FF03 force fields give very different chi(1) rotamer distributions from the coil library. This may partially explain why dipeptide models sometimes cannot reproduce those of protein structures well. The current coil library analysis may be valuable in improving the force field for protein simulations.

Improving Molecular Mechanics Force Fields By Comparison Of Microsecond Simulations With Nmr Experiments

Molecular dynamics simulations and NMR spectroscopy provide complementary approaches to the study of protein structure and dynamics. We have carried out several molecular dynamics simulations of globular proteins and compared the results to a range of NMR experiments that probe the structure and dynamics of these proteins. In particular, simulations on the microsecond timescale allow full sampling of the rotamer distribution of most of the protein side chains. Comparisons with NMR data suggest that, for some residues, this distribution may be incorrectly reproduced by common force fields. We quantified these discrepancies by performing simulations of small helical peptides and comparing the side-chain rotamer distributions with those found in the Protein Data Bank. The potentially problematic residues identified with this procedure were corrected by suitable modification of the force field terms. The performance of the modified force field was evaluated against NMR spectroscopy data.

Side-chain rotamers in protein α-α hairpins and a mechanism of their selection

Several regularities were observed for the distribution of side-chain rotamers in α-α hairpins of globular proteins. In left-turned α-α hairpins, most side chains adopt t rotamers in d-positions and g− rotamers in g-positions. In right-turned α-α hairpins, most side-chains adopt g− rotamers in a-positions and t rotamers in e-positions. Analysis of these regularities suggested that selection of the side-chain conformation in α-α hairpins depends on two main factors, the mode of α-helix packing and the positions of side chains in α-helices. The regularities were explained by the squeezing mechanism: interhelical interactions bring the α-helices close together so that the side chains are squeezed out of the helix-helix interface and adopt unique conformations.

Influence of solvents and leucine configuration at position 5 on tryptophan fluorescence in cyclic enkephalin analogues.

The fluorescence decay of tryptophan is a sensitive indicator of its local environment within a peptide or protein. In this study we carried out fluorescence measurements of the tryptophan residue of cyclic enkephalin analogues of a general formula X-c[D-Dab(2)-Gly(3)-Trp(4)-Y(5)] where X = Cbz or H and Y = D- or L-Leu, in four solvents [water, methanol, acetonitrile, and dimethyl sulfoxide (DMSO)]. An analysis of the tryptophan fluorescence decays using a discrete-exponential model indicates that tryptophan fluorescence decay can be described by a double exponential function in all solvents studied. Lifetime distribution analysis yields a bimodal distribution in protic solvents (water and methanol), whereas an asymmetric, unimodal distribution in an aprotic solvent (DMSO) and uni- or bimodal distributions in acetonitrile solution, depending on leucine configuration. The data are interpreted in terms of the rotamer model, in which the modality and the relative proportions of the lifetime components are related to the population distribution of tryptophan chi(1) rotamers about the C(alpha)--C(beta) bond. The chirality of the Leu(5) residue and solvent properties affect the local environment of the tryptophan residue and therefore influence the distribution of side-chain rotamers. These results are consistent with the results of theoretical conformational calculations.

Tryptophan Rotamer Distributions in Amphipathic Peptides at a Lipid Surface

The fluorescence decay of tryptophan is a sensitive indicator of its local environment within a peptide or protein. We describe the use of frequency domain fluorescence spectroscopy to determine the conformational and environmental changes associated with the interaction of single tryptophan amphipathic peptides with a phospholipid surface. The five 18-residue peptides studied are based on a class A amphipathic peptide known to associate with lipid bilayers. The peptides contain a single tryptophan located at positions 2, 3, 7, 12, or 14 in the sequence. In aqueous solution, the peptides are unstructured and a triple-exponential function is required to fit the decay data. Association of the peptides with small unilamellar vesicles composed of egg phosphatidylcholine reduces the complexity of the fluorescence decays to a double exponential function, with a reduced dependence of the preexponential amplitude on peptide sequence. The data are interpreted in terms of a rotamer model in which the modality and relative proportions of the lifetime components are related to the population distribution of tryptophan χ 1 rotamers about the C α -C β bond. Peptide secondary structure and the disposition of the tryptophan residue relative to the lipid and aqueous phases in the peptide-lipid complex affect the local environment of tryptophan and influence the distribution of side-chain rotamers. The results show that measurement of the temporal decay of tryptophan emission provides a useful adjunct to other biophysical techniques for investigating peptide-lipid and protein-membrane interactions.

Analysis of Thermodynamic Determinants in Helix Propensities of Nonpolar Amino Acids through a Novel Free Energy Calculation

The relative helix propensities of Gly, Ala, Val, Ile, and Leu in the center of a polyalanine helix were calculated using a novel free energy simulation method (Wang et al. J. Mol. Biol. 1995, 253, 473) that permits the decomposition of the free energy into its various thermodynamic components. The calculated relative free energy changes agree well with the recent set of experimental data of Chakrabartty et al. (Protein Science 1994, 3, 843) on alanine-based peptides. The side chain rotamer distributions in the α-helix produced are also consistent with the reports in the literature based on a statistical survey of crystal structures of proteins. A detailed decomposition of the free energy showed that the solvation effect, or hydrophobicity in particular, has little contribution to the helix propensities of the amino acids relative to Gly. The side chain−helical matrix van der Waals interactions are generally favorable and account for a large part of the free energy change relative to Gly upon helix foldin...

α‐Helix‐forming propensities in peptides and proteins

Much effort has been invested in seeking to understand the thermodynamic basis of helix stability in both peptides and proteins. Recently, several groups have measured the helix-forming propensities of individual residues (Lyu, P.C., Liff, M.I., Marky, L.A., Kallenbach, N.R. Science 250:669-673, 1990; O'Neil, K.T., DeGrado, W.F. Science 250:646-651, 1990; Padmanabhan, S., Marqusee, S., Ridgeway, T., Laue, T.M., Baldwin, R.L. Nature (London) 344:268-270, 1990). Using Monte Carlo computer simulations, we tested the hypothesis that these differences in measured helix-forming propensity are due primarily to loss of side chain conformational entropy upon helix formation (Creamer, T.P., Rose, G.D. Proc. Natl. Acad. Sci. U.S.A. 89:5937-5941, 1992). Our previous study employed a rigid helix backbone, which is here generalized to a completely flexible helix model in order to ensure that earlier results were not a methodological artifact. Using this flexible model, side chain rotamer distributions and entropy losses are calculated and shown to agree with those obtained earlier. We note that the side chain conformational entropy calculated for Trp in our previous study was in error; a corrected value is presented. Extending earlier work, calculated entropy losses are found to correlate strongly with recent helix propensity scales derived from substitutions made within protein helices (Horovitz, A., Matthews, J.M., Fersht, A.R. J. Mol. Biol. 227:560-568, 1992; Blaber, M., Zhang, X.-J., Matthews, B.M. Science 260:1637-1640, 1993). In contrast, little correlation is found between these helix propensity scales and the accessible surface area buried upon formation of a model polyalanyl alpha-helix. Taken in sum, our results indicate that loss of side chain entropy is a major determinant of the helix-forming tendency of residues in both peptide and protein helices.

Nuclear magnetic resonance conformational studies on the chemotactic tripeptide formyl-L-methionyl-L-leucyl-L-phenylalanine. A small beta sheet.

Previous work by several groups has shown that the combination of spin--spin coupling constants and spectral density components (derived from spin--lattice relaxation and/or nuclear Overhauser measurements) may aid in the task of conformational determination of peptides in solution. Using the peptide formyl-L-methionyl-L-leucyl-L-phenylalanine, which is a potent specific chemotactic agent for leucocytes, we show the following: (a) that 3JNHCH coupling constants are consistent with a high degree of rigidity in the peptide backbone in solution, (b) that 3H isotopic substitution in combination with relaxation data taken at different Larmor frequencies enables spectral density, and thence conformational, information to be obtained, (c) that side-chain conformations for this molecule mirror, in some aspects, those found in the solid state for other peptides containing the same residues, and (d) that temperature dependence of amide chemical shifts does not have direct implication concerning the existence of intramolecular hydrogen bonds in peptides. We are able to propose a family of conformations which appear to interchange rapidly on the NMR time scale and are characterized by a distribution of side-chain rotamers. The basic backbone conformation is, or closely approximates, a small beta antiparallel pleated sheet and as such suggests a possible mode of receptor--chemotactic peptide interaction.