Coarse Grained Models Reveal Essential Contributions of Topological Constraints to the Conformational Free Energy of RNA Bulges

[Image: see text] Recent studies have shown that simple stereochemical constraints encoded at the RNA secondary structure level significantly restrict the orientation of RNA helices across two-way junctions and yield physically reasonable distributions of RNA 3D conformations. Here we develop a new coarse-grain model, TOPRNA, that is optimized for exploring detailed aspects of these topological constraints in complex RNA systems. Unlike prior models, TOPRNA effectively treats RNAs as collections of semirigid helices linked by freely rotatable single strands, allowing us to isolate the effects of secondary structure connectivity and sterics on 3D structure. Simulations of bulge junctions show that TOPRNA captures new aspects of topological constraints, including variations arising from deviations in local A-form structure, translational displacements of the helices, and stereochemical constraints imposed by bulge-linker nucleotides. Notably, these aspects of topological constraints define free energy landscapes that coincide with the distribution of bulge conformations in the PDB. Our simulations also quantitatively reproduce NMR RDC measurements made on HIV-1 TAR at low salt concentrations, although not for different TAR mutants or at high salt concentrations. Our results confirm that topological constraints are an important determinant of bulge conformation and dynamics and demonstrate the utility of TOPRNA for studying the topological constraints of complex RNAs.