Global analysis of riboswitches by small-angle X-ray scattering and calorimetry

Riboswitches are phylogenetically widespread non-coding mRNA domains that directly bind cellular metabolites and regulate transcription, translation, RNA stability or splicing via alternative RNA structures modulated by ligand binding. The details of ligand recognition by many riboswitches have been elucidated using X-ray crystallography and NMR. However, the global dynamics of riboswitch-ligand interactions and their thermodynamic driving forces are less understood. By compiling the work of many laboratories investigating riboswitches using small-angle X-ray scattering (SAXS) and isothermal titration calorimetry (ITC), we uncover general trends and common themes. There is a pressing need for community-wide consensus experimental conditions to allow results of riboswitch studies to be compared rigorously. Nonetheless, our meta-analysis reveals considerable diversity in the extent to which ligand binding reorganizes global riboswitch structures. It also demonstrates a wide spectrum of enthalpy-entropy compensation regimes across riboswitches that bind a diverse set of ligands, giving rise to a relatively narrow range of physiologically relevant free energies and ligand affinities. From the strongly entropy-driven binding of glycine to the predominantly enthalpy-driven binding of c-di-GMP to their respective riboswitches, these distinct thermodynamic signatures reflect the versatile strategies employed by RNA to adapt to the chemical natures of diverse ligands. Riboswitches have evolved to use a combination of long-range tertiary interactions, conformational selection, and induced fit to work with distinct ligand structure, charge, and solvation properties. This article is part of a Special Issue entitled: Riboswitches.