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...

Solvation of a tetrapeptide, NAc-Ser-Phe-Val-Gly-OMe (1), in water and in water/alcohol mixtures with 2,2,2-trifluoroethanol (TFE)/water or ethanol (ETH)/water has been studied by diffusion NMR and intermolecular NOE measurements. The experimental results were compared with those obtained from detailed Molecular Dynamics (MD) calculations. Independently, all three methods revealed preferential solvation on the surface of the peptide by TFE in the water/TFE mixtures, but not by ETH in the water/ETH mixtures. The MD calculations show that the TFE concentration coating the peptide is higher than that in the bulk, while for ethanol, the concentration is nearly equal to that in the bulk. Calculated site-specific preferential solvation data between TFE, ETH, and water with the different peptide groups have been compared with the NMR data and shown to be in general agreement with the experimental facts.

An all-atom model of 1,1,1,3,3,3-hexafluoro-propan-2-ol (HFIP) for use in molecular dynamics simulation studies is proposed. The model was parametrized by fitting to the experimental density, pressure, and enthalpy of vaporization of the pure liquid at 298 K. The model was then tested by comparison against other experimental thermodynamic and kinetic properties of the pure liquid. Mixtures with SPC water were also investigated. Overall, reasonable agreement with the available experimental data for the neat liquid and for mixtures with SPC water was found. A tendency for HFIP to cluster in SPC water was observed in qualitative agreement with experimental observations. The effect of HFIP on the secondary structure of peptides was also studied. Two simulations of the peptide Melittin, in pure water and in 30% v/v HFIP, demonstrate the helix stabilizing effect of HFIP.

A new model for 2,2,2-trifluoroethanol is proposed. It is a 7-atom model with the methylene group treated as an united atom. The model was optimized to reproduce the physicochemical properties of the pure liquid. The properties of the new model were compared with the available experimental data over a range of temperatures. Furthermore, mixtures with the SPC water model were simulated to assess the ability to reproduce available thermodynamic and kinetic data as well as dielectric properties. The model provides a good agreement with experimental data for the neat liquid and for mixtures with water.

The molecular mechanism by which HFIP stabilizes the a-helical structure of peptides is not well understood. In the present study, we use melittin as a model to gain insight into the details of the atomistic interactions of HFIP with the peptide. We have performed extensive comparative molecular dynamics simulations (up to 100 nsec) in the absence and in the presence of HFIP. In agreement with recent NMR experiments, the simulations show rapid loss of tertiary structure in water at pH 2 but much higher helicity in 35% HFIP. The MD simulations also indicate that melittin adopts a highly dynamic global structure in 35% HFIP solution with two a-helical segments sampling a wide range of angular orientations. The analysis of the HFIP distribution shows the tendency of HFIP to aggregate around the peptide, increasing the local cosolvent concentration to more than two times that in the bulk concentration. The correlation of local peptide structure with HFIP coating suggests that displacement of water at the peptide surface is the main contribution of HFIP in stabilizing the secondary structure of melittin. Finally, a stabilizing effect promoted by the presence of counter-ions was also observed in the simulations.