Osmolyte structural and thermodynamic effects across the protein folding landscape

In this work, the structure and thermodynamics of urea and TMAO preferential interactions across the complete folding landscape of the β-sheet SH3 domain and the helical B domain of protein A (BdpA) are characterized. There is a high correlation between preferential interactions and the surface area of the denatured states, despite the fact that the chemical nature of the exposed surfaces upon denaturation differs from those of native states. For SH3, the denaturation always proceeds with an increase in surface area, such that the qualitative effect of cosolvents on the stability of all denatured states can be inferred from the preferential interactions in the native state. On the other hand, for BdpA, partially denatured states exist that are destabilized by urea; thus, unfolding pathways can be modulated by the cosolvent. Both urea and TMAO form hydrogen bonds with the proteins, which are weakened upon denaturation, and nonspecific interactions, which are strengthened in unfolded structures. Backbone, side chain, and specific residue contributions to distribution functions are obtained, illustrating, for instance, the crucial participation of urea-backbone and nonpolar-cosolvent interactions in the solvation mechanisms. Obtaining these results was possible using a novel computational pipeline to represent solvation structures throughout complete folding landscapes by means of coarse-grained and atomistic simulations and, crucially, the analysis of solvation using minimum-distance distribution functions and the Kirkwood-Buff solvation theory. Cosolvent effects on transfer free energies match experimental data within 1 kcal mol ^-1 , supporting the nuanced description of the osmolyte-protein interplay. The proposed methods can be applied to the study of solvation structure and thermodynamics in other complex molecular systems undergoing large conformational variations, such as non-biological macromolecules and aggregates.