Phase Separation Modulates the Thermodynamics and Kinetics of RNA Hybridization

Biomolecular condensates can influence cellular function in a number of ways, including by changing the structural dynamics and conformational equilibria of the molecules partitioned within them. Here we use methyl transverse relaxation optimized spectroscopy (methyl-TROSY) NMR in conjunction with 2'-O-methyl labeling of RNA to characterize the thermodynamics and kinetics of RNA-RNA base pairing in condensates formed by the C-terminal intrinsically disordered region of CAPRIN1, an RNA-binding protein involved in RNA transport, translation, and stability. CAPRIN1 condensates destabilize RNA-RNA base pairing, resulting from a ∼270-fold decrease and a concomitant ∼15-fold increase in the on- and off-rates for duplex formation, respectively. The ∼30-fold slower diffusion of RNA single strands within the condensed phase partially accounts for the reduced on-rate, but the further ∼9-fold reduction likely reflects shedding of CAPRIN1 chains that are interacting with the RNA prior to hybridization. Our study emphasizes the important role of protein solvation in modulating nucleic acid recognition processes inside condensates.