Destabilization of Hydrophobic Core of Chicken Villin Headpiece in Guanidinium Chloride Induced Denaturation: Hint of π-Cation Interaction.

Abstract

Despite their routine use as protein denaturants, the comprehensive understanding of the molecular mechanisms by which urea and guanidinium chloride (GdmCl) disrupts proteins' structure is still lacking. Here, we use steered molecular dynamics simulations along with the umbrella sampling technique to elucidate the mechanism of unfolding of chicken villin headpiece (HP-36) in these two denaturants. We find that while urea denatures protein predominantly by forming hydrogen bonds with the protein backbone, GdmCl commences unfolding by weakening of the hydrophobic interactions present in the core. The potential of mean force calculation indicates the reduction of hydrophobic interactions between two benzene moieties in 6 M GdmCl as compared to 6 M urea. We observe a near parallel orientation between the guanidinium cation and aromatic side chains of the HP-36 suggesting π-cation type stacking interactions which play a crucial role in weakening of the hydrophobic interaction. We use QM/MM optimization calculations to estimate the energetics of this π-cation interaction. Additionally, the consistency of the unfolding paths between high temperature (400 K) unfolding simulations and steered molecular dynamics simulations strengthens the proposed molecular mechanism of unfolding further.