Experimental and Computational Studies of Liquid-Liquid Phase Separation of Some Globular Proteins

Abstract

Proteins are complex biomolecules that are involved in everything that enables the normal functioning of all living organisms. Consequently, even their minor change or instability can have devastating consequences for life itself. Over the last decade, liquid-liquid phase separation (LLPS) of proteins has been associated with the onset of numerous diseases, therefore gained knowledge in this field is of vital importance for disease prevention and treatment. Despite thorough research work on LLPS of proteins, many open questions remain to be resolved. In recent years, our studies on LLPS of globular proteins were carried out as an interplay of various experimental techniques, theoretical approach as well as computer simulations. Buffer identity is often neglected, but our results indicate it is non-negligible when discussing the phase stability of protein solutions. At the same time, we managed to extend our coarse-grained protein model to incorporate buffer-specific effects into LLPS theoretical predictions. Through experiments, supported by molecular dynamics simulations, we have demonstrated that arginine can affect the phase stability of protein solutions as a double-edged sword. It can reduce protein self-association when added as free arginine molecules in solution and enhance it when being protein-bound as residues on protein surface. By means of mutations of ARG with LYS on protein surface, we have shed light on the importance of hydrophobic interactions for the protein self-assembly.