Principles of Ligand Modulation of Phase Behavior in Multicomponent Systems

Abstract

Many biomolecular condensates are thought to form via spontaneous phase transitions. A typical condensate encompasses hundreds of distinct types of biomolecules. Accordingly, the components of condensates are classified as scaffolds vs. clients. However, so-called clients do not just partition passively into condensates formed by scaffolds. Instead, they can actually modulate the phase behavior of scaffolds. Here, we coopt the theory of binding and linkage and separate clients into two categories namely, ligands and crowders. We show that preferential binding of ligands to scaffold molecules in different phases will lead to a modulation of phase boundaries via a phenomenon known as polyphasic linkage. Next, we generalize the theory of polyphasic linkage to include the effects of multiple competing ligands. This analysis shows that a network of ligands can enable control over condensates through regulation of the expression levels and preferential interactions of ligands. Further generalizations help us query the effects of preferential binding of ligands on the full coexistence curve and the critical point of a scaffold molecule. Our formalism also allows us to distinguish bona fide crowders from ligands. Crowders alter the free volume of dilute phases and drive phase transitions via depletion-mediated attractions. This distinction allows us to establish a set of guidelines to help delineate the effects of ligands vs. crowders on the phase behavior of scaffolds. We reevaluate extant data regarding the effects of polyethylene glycol (PEG) and show that in many cases PEG acts as a preferentially interacting ligand rather than a crowder. Our studies have broad implications for the use of synthetic crowders in vitro and for understanding the effects of ligands on phase behavior in vivo.