Structural transitions and thermodynamics of a glycine-dependent riboswitch from Vibrio cholerae

J Mol Biol. 2007 Feb 2;365(5):1393-406. doi: 10.1016/j.jmb.2006.10.022. Epub 2006 Oct 13.


Riboswitches are complex folded RNA domains found in noncoding regions of mRNA that regulate gene expression upon small molecule binding. Recently, Breaker and coworkers reported a tandem aptamer riboswitch (VCI-II) that binds glycine cooperatively. Here, we use hydroxyl radical footprinting and small-angle X-ray scattering (SAXS) to study the conformations of this tandem aptamer as a function of Mg(2+) and glycine concentration. We fit a simple three-state thermodynamic model that describes the energetic coupling between magnesium-induced folding and glycine binding. Furthermore, we characterize the structural conformations of each of the three states: In low salt with no magnesium present, the VCI-II construct has an extended overall conformation, presumably representing unfolded structures. Addition of millimolar concentrations of Mg(2+) in the absence of glycine leads to a significant compaction and partial folding as judged by hydroxyl radical protections. In the presence of millimolar Mg(2+) concentrations, the tandem aptamer binds glycine cooperatively. The glycine binding transition involves a further compaction, additional tertiary packing interactions and further uptake of magnesium ions relative to the state in high Mg(2+) but no glycine. Employing density reconstruction algorithms, we obtain low resolution 3-D structures for all three states from the SAXS measurements. These data provide a first glimpse into the structural conformations of the VCI-II aptamer, establish rigorous constraints for further modeling, and provide a framework for future mechanistic studies.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Aptamers, Nucleotide / chemistry*
  • Aptamers, Nucleotide / genetics
  • Base Sequence
  • Glycine / metabolism*
  • Hydroxyl Radical / metabolism
  • Magnesium / pharmacology
  • Models, Molecular
  • Molecular Sequence Data
  • Nucleic Acid Conformation / drug effects
  • Nucleic Acid Denaturation / drug effects
  • RNA, Bacterial / chemistry*
  • RNA, Bacterial / genetics
  • Scattering, Small Angle
  • Solvents
  • Thermodynamics
  • Vibrio cholerae / chemistry*
  • X-Ray Diffraction


  • Aptamers, Nucleotide
  • RNA, Bacterial
  • Solvents
  • Hydroxyl Radical
  • Magnesium
  • Glycine