Single transcriptional and translational preQ1 riboswitches adopt similar pre-folded ensembles that follow distinct folding pathways into the same ligand-bound structure

Authors

Krishna C. Suddala, University of Michigan - Ann Arbor
Arlie J. Rinaldi, University of Michigan - Ann Arbor
Jun Feng, University of Michigan - Ann Arbor
Anthony M. Mustoe, University of Michigan - Ann Arbor
Catherine D. Eichhorn, University of Michigan - Ann ArborFollow
Joseph A. Liberman, University of Rochester School of Medicine and Dentistry
Joseph E. Wedekind, University of Rochester School of Medicine and Dentistry
Hashim M. Al-Hashimi, University of Michigan - Ann Arbor
Charles L. Brooks III, University of Michigan - Ann Arbor
Nils G. Walter, University of Michigan - Ann ArborFollow

Document Type

Article

Date of this Version

2013

Citation

Nucleic Acids Research, 2013, Vol. 41, No. 22

Comments

© The Author(s) 2013.

Open access

doi:10.1093/nar/gkt798

Abstract

Riboswitches are structural elements in the 50 untranslated regions of many bacterial messenger RNAs that regulate gene expression in response to changing metabolite concentrations by inhibition of either transcription or translation initiation. The preQ₁ (7-aminomethyl-7-deazaguanine) riboswitch family comprises some of the smallest metabolite sensing RNAs found in nature. Once ligand-bound, the transcriptional Bacillus subtilis and translational Thermoanaerobacter tengcongensis preQ₁ riboswitch aptamers are structurally similar RNA pseudoknots; yet, prior structural studies have characterized their ligand-free conformations as largely unfolded and folded, respectively. In contrast, through single molecule observation, we now show that, at nearphysiological Mg²⁺ concentration and pH, both ligand-free aptamers adopt similar pre-folded state ensembles that differ in their ligand-mediated folding. Structure-based Go¯ -model simulations of the two aptamers suggest that the ligand binds late (Bacillus subtilis) and early (Thermoanaerobacter tengcongensis) relative to pseudoknot folding, leading to the proposal that the principal distinction between the two riboswitches lies in their relative tendencies to fold via mechanisms of conformational selection and induced fit, respectively. These mechanistic insights are put to the test by rationally designing a single nucleotide swap distal from the ligand binding pocket that we find to predictably control the aptamers0 pre-folded states and their ligand binding affinities.