Enhancing Protein Stability: Tuning Hydrophobic Interactions with Additives

Among the various forces involved in protein folding and stability, hydrophobic interactions are dominant contributors. Introduction of additives to solutions can be used to modulate the strength of hydrophobic interactions, and in turn, the stability of proteins. Here, we investigated the effects of several amino acids and sugars – which are common additives in protein formulations – on the folding and stability of a model hydrophobic polymer. We show that amino acids and sugars modulate hydrophobic interactions through a combination of direct and indirect mechanisms, depending on their concentration. We also find that the amino acid side chain and polar tail play different roles in driving these mechanisms and the nature of the side chain determines the mechanism of action. We demonstrate that the complex and context-dependent effects provided by these amino acids and sugars on protein stability can be explained by the balance between direct and indirect contributions. This work provides insights into the molecular-level mechanisms of additive-induced stabilization and lays the foundation for future studies on rational formulation design.