Ligand Recognition And Conformational Dynamics Of Sam-Binding Riboswitches

Abstract

Riboswitches are recently discovered regulatory RNAs that directly relay environmental cues to the genetic regulat ory machinery. S box (SAM-I) and the SMK box (SAM-III) RNA are both S- Adenosyl -L-methionine (SAM) responsive riboswitches that regulate bacterial gene expression at the levels of transcription attenuation or translation inhibition respectively. The S box RNA is the most wide spread transcriptional riboswitch in Gram -positive bacteria, while the SMK RNA is a translational riboswitch in members of the Lactobacillales. Both RNAs bind specifically to SAM and feedback regulate the SAM biosynthetic pathway. However, these riboswitches share no sequence, structural or mechanistic homology. Our lab employs both X-ray crystallography and chemical probing to analyze their ligand recognition and mechanisms of gene repression. Our X-ray structures show that both the SMK and S box riboswitches fold into compact structures once bound to SAM, and that both RNAs adopt unique yet elegantly simple ways to specifically recognize SAM in favor of its close homolog, SAdenosyl-homocysteine (SAH). Further mutagenesis and chemical probing results show that both the SMK and S box RNA adopt extremely dynamic conformations in the absence of SAM, and certain structural elements outside the SAM-binding domain have profound effects on the conformation dynamics. The thermal melting experiments also showed that both riboswitches undergo unique unfolding processes when subject to increasing temperature s. Our findings suggest that these SA