Solvent effects on protein folding thermodynamics

Protein structures and their folding mechanisms are dependent on the molecular environment. Proteins can adopt distinct conformations – active, inactive, or with different functionalities – depending on temperature, pressure, or the characteristics of the solvent in which they are immersed. Although proteins perform their functions primarily in aqueous environments, even within the cellular milieu, various species, such as cosolvents and ions, influence their structure and function. Additionally, some cosolvents are used in vitro to stabilize proteins and enhance their function in biotechnological applications. Cosolvents are present in living organisms and various formulations, but the mechanism by which they affect protein stability – whether in the native state or non-native conformations – is often unclear. This thesis explores protein folding and how different solvents affect its thermodynamics. In Chapter 3, we investigate the folding of the (AAQAA)3 peptide in water and 2,2,2-Trifluoroethanol (TFE) solutions to elucidate TFE’s stabilizing mechanisms. We employ Replica Exchange Molecular Dynamics simulations to enhance peptide sampling. In Chapter 4, we analyze how the folding of the B domain of protein A (BdpA) is associated with its secondary structure formation and hydration structures. In this case, we use simulations with Structure-Based Models (SBMs). Our results allowed us to group partially folded BdpA structures, enabling an understanding of the effects of osmolytes on the relative stability of folding ensembles, which are discussed in detail in Chapter 5. Chapter 5 examines how the osmolytes urea and TMAO contribute to the stabilization mechanisms of different folding ensembles of the SH3 domain and BdpA, proteins with distinct structural motifs widely adopted as models for folding studies. Solvent effects are analyzed using Minimum-Distance Distribution Functions (MDDFs) and Kirkwood-Buff theory. Finally, in Chapter 6, we address aspects of the folding mechanism of the MAD2 protein (Mitotic Spindle Assembly Checkpoint Protein). MAD2 undergoes reversible folding into two distinct native states: inactive and active. The model developed in this work captures the transition between these two states.