Structural analysis of a class III preQ sub( 1) riboswitch reveals an aptamer distant from a ribosome-binding site regulated by fast dynamics

PreQ...-III riboswitches are newly identified RNA elements that control bacterial genes in response to preQ... (7-aminomethyl-7-deazaguanine), a precursor to the essential hypermodified tRNA base queuosine. Although numerous riboswitches fold as H-type or HL...-type pseudoknots that integrate ligand-binding and regulatory sequences within a single folded domain, the preQ...-III riboswitch aptamer forms a HL...-type pseudoknot that does not appear to incorporate its ribosome-binding site (RBS). To understand how this unusual organization confers function, we determined the crystal structure of the class III preQ... riboswitch from Faecalibacterium prausnitzii at 2.75 ... resolution. PreQ1 binds tightly (K... 6.5 plus or minus 0.5 nM) between helices P1 and P2 of a three-way helical junction wherein the third helix, P4, projects orthogonally from the ligand-binding pocket, exposing its stem-loop to base pair with the 3' RBS. Biochemical analysis, computational modeling, and single-molecule FRET imaging demonstrated that preQ... enhances P4 reorientation toward P1-P2, promoting a partially nested, H-type pseudoknot in which the RBS undergoes rapid docking (k... ~0.6 s...) and undocking (k... ~1.1 s...). Discovery of such dynamic conformational switching provides insight into how a riboswitch with bipartite architecture uses dynamics to modulate expression platform accessibility, thus expanding the known repertoire of gene control strategies used by regulatory RNAs. (ProQuest: ... denotes formulae/symbols omitted.)