Environmental influence on alpha-helical peptides

Abstract

Most proteins at physiological conditions fold into a native functional three-dimensional conformation. The stability of the Native state is a balance between different interactions within the protein and with the solvent. The role of the solvent is therefore important for the stability of proteins and determinant for their three-dimensional structure and function.

Secondary structure elements such as a helices are often found in the nucleation core structures proposed by the nucleation-diffusion model for protein folding. Understanding their stability and their folding/unfolding mechanism can shed light on the protein folding problem.

This thesis is devoted to the study of α-helical peptides in different environments, where they show very different behavior.

Proteins unfold at high denaturant concentrations, such as urea or guanidinium. Their solubility increases with denaturant concentration. The particular properties of the urea-aqueous solutions are still not well understood. Urea dimerization has been pointed out as a fundamental factor for the thermodynamical behavior of urea-water solutions. In the first work, we calculate urea molecule parameters consistent with the CHARMM force field and TIP3 water model taking in account the urea dimerization feature. Simulations of 2M and 8M urea contained significant amount of cyclic dimers with very favorable interactions as well as small amount of head-to-tail dimers, as found in crystals Comparisons with potentials obtained from ab initio calculations indicate the non adequacy of the parameters obtained with this methodology. Furthermore, the DFT results indicate that urea may adopt a planar conformation in solution as it does in crystals.

The urea parameters previously obtained were used to perform MD simulations of two peptides: a C-peptide analogue and a variant (same amino acid composition but different sequence) in 8M urea-water solution and in water at different temperatures. The results indicated that the stability of the C-peptide arises from electrostatic side chain-side chain interactions. In contrast, the second peptide was unstable in all solvents. In the presence of urea the ability of water molecules to form hydrogen bonds to the peptide as well as their life time is increased. The rotation of the water molecules is reduced on the peptide surface. In addition, urea formed long lived hydrogen bonds to the peptide and accumulated in excess on the peptide surface.

MD simulations of the putative α-helical transmembrane (TM) spanning domain of two glycoproteins E1 and E2 of the Semliki Forest virus wild type and mutants were performed. The two helices pack in left handed two-stranded rope fashion. The residues that occupy the important positions of the heptad (abcdefg) were identified. Success of packing was a compromise between small and medium residues at E1 and E2 interfaces that allow hydrogen bond formation. Defects on packing were predicted. The method can be extrapolated to sketch TM helical packing in other alphaviruses.

The dynamical properties of the peptide hormone motilin (22 amino acids) were studied. The MD simulations in water and HFIP coincide with the NMR and fluorescence anisotropy decay (FAD) results. However, the choice of the vectors under study is very important to get agreement. NMR relaxation parameters and spectral density functions were also obtained from the simulations which evidenced the α-helical character of the peptide but also its high flexibility even in the HFIP-water solution. The results, point out the difficulty for using NMR dynamical analysis with flexible peptides.