Dissecting urea-RNA interactions in SARS-CoV-2 virus: nucleobases as the primary denaturation hotspots.

RNA structure function relationships are of particular significance in the context of the SARS-CoV-2 genome, yet the molecular principles governing chemical denaturation of large, structured RNAs remain elusive. Here, we use all-atom molecular dynamics simulations to resolve how aqueous urea unfolds the SARS-CoV-2 frameshift stimulation element (FSE), a conserved RNA motif essential for viral translation. We show that urea induces RNA unfolding through a nucleobase-selective mechanism, rather than nonspecific backbone solvation. Urea preferentially concentrates at solvent-exposed bases, where it forms directional hydrogen bonds and stacking interactions that compete with native base pairing and expel interfacial water. This targeted solvation increases base mobility, amplifies conformational heterogeneity, and progressively destabilizes the intramolecular hydrogen-bond network. The denaturation process is marked by a sigmoidal transition in the unfolded RNA population and reshaping of the free-energy landscape at surprisingly low to higher urea concentrations. These results identify nucleobase solvation as the dominant driving force for urea-induced RNA unfolding and establish a general physicochemical framework for chemical denaturation of functional viral RNAs.