Probing the Structural Determinants of Type II‘ β-Turn Formation in Peptides and Proteins

Gibbs, Alan C.; Bjorndahl, Trent C.; Hodges, Robert S.; Wishart, David S.

doi:10.1021/ja011005e

Cited by 91 publications

(107 citation statements)

References 62 publications

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“…Tyr49 seems to provide stabilizing interactions at this position as a Y49N mutation causes a decrease in stability of 1 kcal/mol (ST, unpublished data). These results as well as the results of Gibbs et al 41 suggest that side chains with aromatic rings can provide stabilizing interactions at the i+1 position of a type II turn.…”

Section: Resultssupporting

confidence: 71%

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Amino Acid Contribution to Protein Solubility: Asp, Glu, and Ser Contribute more Favorably than the other Hydrophilic Amino Acids in RNase Sa

Treviño

Scholtz

Pace

2007

Journal of Molecular Biology

209

178

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SUMMARYPoor protein solubility is a common problem in high resolution structural studies, formulation of protein pharmaceuticals, and biochemical characterization of proteins. One popular strategy to improve protein solubility is to use site-directed mutagenesis to make hydrophobic to hydrophilic mutations on the protein surface. However, a systematic investigation of the relative contributions of all twenty amino acids to protein solubility has not been done. Here, twenty variants at the completely solvent-exposed position 76 of Ribonuclease (RNase) Sa are made to compare the contributions of each amino acid. Stability measurements were also made for these variants, which occur at the i+1 position of a type II β-turn. Solubility measurements in ammonium sulfate solutions were made at high positive net charge, low net charge, and high negative net charge. Surprisingly, there was a wide range of contributions to protein solubility even among the hydrophilic amino acids. The results suggest that aspartic acid, glutamic acid, and serine contribute significantly more favorably than the other hydrophilic amino acids especially at high net charge. Therefore, to increase protein solubility, asparagine, glutamine, or threonine should be replaced with aspartic acid, glutamic acid or serine.

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Section: Resultssupporting

confidence: 71%

“…Gibbs et. al 41 investigated the role of amino acid substitutions in type II' β-turns. Type II' β-turns are mirror images of type II β-turns.…”

Section: Resultsmentioning

confidence: 99%

Amino Acid Contribution to Protein Solubility: Asp, Glu, and Ser Contribute more Favorably than the other Hydrophilic Amino Acids in RNase Sa

Treviño

Scholtz

Pace

2007

Journal of Molecular Biology

209

178

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“…In fact the predominate effect of TFE on BET and BHA seems to be in the stabilization of the turn regions. The importance of turn regions in determining the folding and the stability of ␤-forming structures has been shown by several groups (35,41,42), and the selective stabilization of turn regions by TFE has been demonstrated by several recent NMR studies (34,35,41), which certainly could in part account for the effect of TFE on the stability ␤-sheet-forming peptides.…”

Section: Discussionmentioning

confidence: 94%

“…Whereas in TFE͞water the geometry of the turn is well conserved throughout the simulation, in water the turn geometry fluctuates being recognized approximately a third of the time as the bend. These fluctuations are in turn related to the twisting of the peptide (9,34,35).…”

Section: Resultsmentioning

confidence: 99%

Mechanism by which 2,2,2-trifluoroethanol/water mixtures stabilize secondary-structure formation in peptides: A molecular dynamics study

Roccatano

Colombo

Fioroni

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

467

452

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Molecular dynamics simulation techniques have been used to investigate the effect of 2,2,2-trifluoroethanol (TFE) as a cosolvent on the stability of three different secondary structure-forming peptides: the ␣-helix from Melittin, the three-stranded ␤-sheet peptide Betanova, and the ␤-hairpin 41-56 from the B1 domain of protein G. The peptides were studied in pure water and 30% (vol͞vol) TFE͞water mixtures at 300 K. The simulations suggest that the stabilizing effect of TFE is induced by the preferential aggregation of TFE molecules around the peptides. This coating displaces water, thereby removing alternative hydrogen-bonding partners and providing a low dielectric environment that favors the formation of intrapeptide hydrogen bonds. Because TFE interacts only weakly with nonpolar residues, hydrophobic interactions within the peptides are not disrupted. As a consequence, TFE promotes stability rather than inducing denaturation. F or more than three decades, 2,2,2-trifluoroethanol (TFE) has been used as a cosolvent for the study of peptides in solution because NMR and CD studies show that the presence of TFE increases the population of ␣-helix and ␤-sheet content in secondary-structure-forming peptides in TFE͞water mixtures (1-3). Despite the effects of TFE having been known for such a long time, the mechanism by which TFE stabilizes secondary structure in peptides is still not clear. One possible explanation is the preferential solvation of the folded state by TFE (3). According to this hypothesis, TFE acts within the context of a preexisting helix-coil equilibrium, and the preferential interaction of TFE with the folded state shifts the equilibrium toward more structured conformations (3). The molecular nature of the TFE-peptide interaction is not clear, however. Alternative mechanisms have also been proposed to explain the stabilizing effect of TFE. In particular, the effect could result from TFE reinforcing hydrogen bonds between carbonyl and amidic NH groups by the removal of water molecules in the proximity of the solute (4) and͞or the lowering of the dielectric constant (1). Furthermore, small-angle x-ray-scattering studies (2) show that TFE forms clusters in water as the concentration of the organic cosolvent is increased, with a maximum at 30% (vol͞vol) TFE. Reiersen and Rees (5) have proposed that such TFE clusters locally assist the folding of secondary-structure elements by providing a solvent matrix that promotes hydrophobic interactions between amino acid side chains. Recent NMR studies involving small peptides in TFE provide some support for this hypothesis (6-8). Each of these mechanisms could explain the stabilization of ␣-helical peptides in solution (1, 3) but not necessarily the stabilization of ␤-structure (1, 2).In recent years molecular dynamics (MD) simulations have been increasingly used to understand the complex conformational equilibria of polypeptides in solution and to predict structural preferences (9-11). In particular, the importance of side-chain interactions in determining pepti...

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“…A reverse turn conformation was chosen, because it provides the appropriate geometry, topological twist and in-register peptide alignment. 23 In addition, a high immune response requires conjugation to a carrier protein, for which we used tetanus toxoid. Thus, a site for conjugation was incorporated in the contra-turn either by a cysteine or by an N 1 -(S-acetylmercaptoacetyl)lysine residue (Lys-SAMA), both of which can be attached to a sulfhydryl-reactive protein.…”

Section: Peptide Designmentioning

confidence: 99%

Immunogenicity of Peptide-vaccine Candidates Predicted by Molecular Dynamics Simulations

Oomen

Hoogerhout

Bonvin

et al. 2003

Journal of Molecular Biology

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We present an in silico, structure-based approach for design and evaluation of conformationally restricted peptide-vaccines. In particular, we designed four cyclic peptides of ten or 11 residues mimicking the crystallographically observed b-turn conformation of a predicted immunodominant loop of PorA from Neisseria meningitidis. Conformational correctness and stability of the peptide designs, as evaluated by molecular dynamics simulations, correctly predicted the immunogenicity of the peptides. We observed a peptide-induced functional antibody response that, remarkably, exceeded the response induced by the native protein in outer membrane vesicles, without losing specificity for related strains. The presented approach offers tools for a priori design and selection of peptide-vaccine candidates with full biological activity. This approach could be widely applicable: to outer membrane proteins of Gram-negative bacteria, and to other epitopes in a large range of pathogens.

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Probing the Structural Determinants of Type II‘ β-Turn Formation in Peptides and Proteins

Cited by 91 publications

References 62 publications

Amino Acid Contribution to Protein Solubility: Asp, Glu, and Ser Contribute more Favorably than the other Hydrophilic Amino Acids in RNase Sa

Amino Acid Contribution to Protein Solubility: Asp, Glu, and Ser Contribute more Favorably than the other Hydrophilic Amino Acids in RNase Sa

Mechanism by which 2,2,2-trifluoroethanol/water mixtures stabilize secondary-structure formation in peptides: A molecular dynamics study

Immunogenicity of Peptide-vaccine Candidates Predicted by Molecular Dynamics Simulations

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