Interplay Between Biomolecular Folding and Liquid-Liquid Phase Separation

Biomolecules can form condensates through liquid-liquid phase separation (LLPS) to perform biological functions. It is well established that intrinsically disordered proteins play essential roles in driving LLPS, and their conformations tend to adopt more extended states within the dense phase. However, for proteins with folded structures, how the folding of individual chains and phase separation mutually influence each other remains unclear. Here, using the nucleocapsid protein 7 (NCp7) of human immunodeficiency virus 1 (HIV-1) and its cognate RNA as a model system, we investigated the coupling between protein folding and phase separation through coarse-grained molecular dynamics simulations. We found that the folding of NCp7 into its native structure, which contains two zinc finger (ZF) motifs, increases the multivalency of protein-RNA interactions and promotes their co-condensation. In turn, phase separation further stabilizes the native structure of the protein by facilitating non-native interactions within the condensates, where biomolecule concentration and molecular crowding are high. Additionally, RNA exhibits a highly dynamic structure, which enables it to simultaneously engage in protein interactions and adopt interchain double-stranded helical conformations, thereby mediating a balance between protein-RNA attraction and RNA-RNA repulsion. The results of this work suggest that tuning the folding and stability of individual biomolecules (e.g., through metal-ion binding) offers an effective route to modulate biomolecular condensation, highlighting a promising strategy for drug design targeting the macroscopic phase behavior of biomolecules.