Elucidation of the Molecular Interaction Network Underlying Full-Length FUS Conformational Transitions and Its Phase Separation Using Atomistic Simulations

Fused in Sarcoma (FUS) is a multidomain nucleic acid binding protein which orchestrates cellular functions such as gene expression, transcription, and DNA repair through liquid-liquid phase separation (LLPS). While crucial to understanding cellular processes, an atomic-level view of the molecular-level interactions associated with full-length (FL) FUS LLPS remains challenging due to its low solubility <i>in vitro</i>. Here, using all-atom (AA) molecular dynamics (MD) simulations, we examined the conformational dynamics and interactions of FL FUS in both dilute and condensed phases. Comparing two modern force fields (FFs)─Amber ff03ws and ff99SBws-STQ, we found that monomer simulation ensembles generated by both FFs exhibited qualitatively similar intramolecular interaction profiles dominated by intrinsically disordered regions (IDRs). While the two folded domains minimally participated in intramolecular interactions, their stabilities significantly influenced the chain dimension and led to discrepancies compared to experimental data for both FFs. We observed that the Amber ff99SBws-STQ coupled with parameters adopted from the Zinc Amber force field (ZAFF) maintained stable folded domains and improved estimates of the chain dimensions. Finally, a microsecond-time scale simulation of FL FUS condensate revealed an extensive network of electrostatic interactions which are strongly correlated with those that modulate the dilute phase conformations. Overall, insights from our AAMD simulations illuminate the interplay between folded domain stability and IDR interactions in modulating protein conformation and phase separation.