Physical Basis of Protein Liquid-Liquid Phase Separation*

Intracellular membraneless organelles, corresponding to the droplet phase upon liquid-liquid phase separation (LLPS) of mixtures of proteins and possibly RNA, mediate myriad cellular functions [Qin & Zhou 2017 Curr Opin Struct Biol]. Cells use a variety of biochemical signals such as expression level and posttranslational modification to regulate droplet formation and dissolution. Our study focuses on elucidating the physical basis of phase behaviors associated with cellular functions of membraneless organelles, using three complementary approaches. First, we use colloids and polymers, respectively, as models for structured and disordered proteins, to investigate both the common basis for protein phase separation and the unique characteristics of structured and disordered proteins in LLPS [Zhou et al 2018 Trends Biochem Sci]. Disordered proteins are characterized by both extensive attraction throughout the sequence and low energetic cost from steric repulsion, contributing to easy observation of phase separation. Second, we use multi-component patchy particles to investigate the wide range of effects of regulatory components on the droplet formation of driver proteins [Nguemaha & Zhou 2018 Sci Rep]. Third, we have developed a powerful computational method called FMAP for determining liquid-liquid phase equilibria [Qin & Zhou 2014 J Chem Theory Comput; 2016 J Phys Chem B]. By using fast Fourier transform to efficiently evaluate protein-protein interactions, FMAP enables an atomistic representation of the protein molecules. Application to g-crystalins reveals how minor variations in amino-acid sequence, similar to those from posttranslational modifications and disease-associated mutations, lead to drastic differences in critical temperature. These studies contribute to both qualitative and quantitative understanding on the phase behaviors of membraneless organelles and their regulation and dysregulation.