Choline chloride as a nano-crowder protects HP-36 from urea-induced denaturation: Insights from Solvent Dynamics and Protein-Solvent interaction

Abstract

Urea at sufficiently high concentration unfolds the secondary structure of proteins leading to denaturation. In contrast, Choline Chloride (ChCl) and urea, in 1:2 molar ratio form a deep eutectic mixture, a liquid at room temperature and protect proteins from denaturation. In order to get a microscopic picture of this phenomenon, we perform extensive all-atom molecular dynamics simulations on a model protein HP-36. Based on our calculation of Kirkwood-Buff integrals, we analyze the relative accumulation of these osmolytes around the protein. Additional insights are drawn from the analyses of the translational and rotational dynamics of solvent molecules and also from the hydrogen bond auto-correlation functions. In the presence of urea, water shows slow subdiffusive dynamics around the protein backbone as a consequence of stronger interaction of water molecules with the backbone atoms. Urea also shows subdiffusive motion. Addition of choline further slows down the dynamics of urea restricting its inclusion around the protein backbone. Adding to this, choline molecules in the first solvation shell of the protein shows the strongest subdiffusive behavior. In other words, ChCl behaves as a nano-crowder by excluding urea from the protein backbone and thereby slowing down the dynamics of the water layer around the protein. This prevents the protein from denaturation and makes it structurally rigid which in turn is supported by the smaller radius of gyration and root mean square deviation values of HP-36 when ChCl is present in the system.