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Structural analysis of a class III preQ₁ riboswitch reveals an aptamer distant from a ribosome-binding site regulated by fast dynamics
- Liberman, Joseph A., Suddala, Krishna C., Aytenfisu, Asaminew, Chan, Dalen, Belashov, Ivan A., Salim, Mohammad, Mathews, David H., Spitale, Robert C., Walter, Nils G., Wedekind, Joseph E.
- Proceedings of the National Academy of Sciences of the United States of America 2015 v.112 no.27 pp. E3485
- crystal structure, genes, image analysis, models, oligonucleotides, regulatory sequences, transfer RNA
- PreQ ₁-III riboswitches are newly identified RNA elements that control bacterial genes in response to preQ ₁ (7-aminomethyl-7-deazaguanine), a precursor to the essential hypermodified tRNA base queuosine. Although numerous riboswitches fold as H-type or HL ₒᵤₜ-type pseudoknots that integrate ligand-binding and regulatory sequences within a single folded domain, the preQ ₁-III riboswitch aptamer forms a HL ₒᵤₜ-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 ₁ riboswitch from Faecalibacterium prausnitzii at 2.75 Å resolution. PreQ ₁ binds tightly ( K D,ₐₚₚ 6.5 ± 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 ₁ enhances P4 reorientation toward P1–P2, promoting a partially nested, H-type pseudoknot in which the RBS undergoes rapid docking ( k dₒcₖ ∼0.6 s ⁻¹) and undocking ( k ᵤₙdₒcₖ ∼1.1 s ⁻¹). 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.