Diversity of structural responses of riboswitches to cognate ligand binding

Riboswitches are gene-regulatory domains of mRNAs that respond to the intracellular concentration of a small molecule metabolites or second messengers. Riboswitches have been found in archaea, bacteria and eukarya. In addition, synthetic riboswitches have been engineered starting with various in vitro-evolved aptamers. Our analysis of ligand recognition by diverse riboswitches employing X-ray crystallography, small-angle X-ray scattering (SAXS), calorimetry, and other techniques reveal a diversity of structural responses. Some, like the catalytic glmS riboswitch, are stably folded prior to ligand binding. Others, like the natural thiamine-pyrophosphate (TPP), preQ₁ and cyclic diguanylate (c-di-GMP) riboswitches and the artificial tetracycline riboswitch exhibit various degrees of ligand-induced compaction and folding. Although, in general, increased structural complexity correlates with smaller ligand-induced folding transitions, the behavior of different riboswitch classes is highly idiosyncratic. Moreover, even riboswitches of the same class can vary. For instance, SAXS studies of TPP riboswitches from E. coli and A. thaliana show that the plant RNA is far less structured in its ligand-free form, even though the crystal structures of the TPP complexes of the two RNAs are nearly identical. This diversity suggests that the structural (and kinetic) response of riboswitches to ligand binding has been fine-tuned to the specific requirements of their genetic loci by evolution.