Stabilizing Effect of Neighboring Disordered RGG Domain on the Folded State of FUS-RRM

Fused in Sarcoma (FUS) is an RNA-binding protein essential for RNA processing, yet its RNA-recognition motif (RRM) is prone to irreversible unfolding and amyloid aggregation, which is associated with the pathogenesis of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Although the isolated RRM is experimentally known to adopt a stable folded structure, its response to long-range interdomain interactions remains poorly understood. In this work, we address this gap by performing rare-event sampling atomistic molecular dynamics simulations of two systems: isolated RRM and RRM with the flanking RGG sequence using multithermal-multiumbrella on-the-fly probability enhanced sampling (MM-OPES). These simulations allow us to characterize the folding landscape of FUS RRM and examine the specific interactions between the RRM and the adjacent RGG region and how they affect the stability of RRM. Our findings reveal that the disordered RGG segment enhances the stability of the folded RRM by forming stabilizing intramolecular contacts that wrap around the domain. This stabilization is driven by increased fractions of the α₁ helix, β₂, β₃, and the KK loop through a network of targeted multivalent contacts between the RGG and RRM residues. This work reveals how a disordered region stabilizes a folded RNA-binding domain, underscoring the importance of disordered-ordered interdomain coupling in shaping the folding landscape of FUS RRM. These results suggest that disruption of such interactions could destabilize the RRM fold and may contribute to misfolding-prone states relevant to FUS dysfunction.