Dissecting electrostatic screening, specific ion binding, and ligand binding in an energetic model for glycine riboswitch folding
- 1Department of Physics, Stanford University, Stanford, California 94305, USA
- 2Kavli Institute of Nanoscience, Delft University of Technology, 2628 CJ Delft, The Netherlands
- 3Department of Applied Physics, Stanford University, Stanford, California 94305, USA
- 4Department of Biochemistry, Stanford University, Stanford, California 94305, USA
- 5Biophysics Program, Stanford University, Stanford, California 94305, USA
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↵6 These authors contributed equally to this work.
Abstract
Riboswitches are gene-regulating RNAs that are usually found in the 5′-untranslated regions of messenger RNA. As the sugar-phosphate backbone of RNA is highly negatively charged, the folding and ligand-binding interactions of riboswitches are strongly dependent on the presence of cations. Using small angle X-ray scattering (SAXS) and hydroxyl radical footprinting, we examined the cation dependence of the different folding stages of the glycine-binding riboswitch from Vibrio cholerae. We found that the partial folding of the tandem aptamer of this riboswitch in the absence of glycine is supported by all tested mono- and divalent ions, suggesting that this transition is mediated by nonspecific electrostatic screening. Poisson–Boltzmann calculations using SAXS-derived low-resolution structural models allowed us to perform an energetic dissection of this process. The results showed that a model with a constant favorable contribution to folding that is opposed by an unfavorable electrostatic term that varies with ion concentration and valency provides a reasonable quantitative description of the observed folding behavior. Glycine binding, on the other hand, requires specific divalent ions binding based on the observation that Mg2+, Ca2+, and Mn2+ facilitated glycine binding, whereas other divalent cations did not. The results provide a case study of how ion-dependent electrostatic relaxation, specific ion binding, and ligand binding can be coupled to shape the energetic landscape of a riboswitch and can begin to be quantitatively dissected.
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Footnotes
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Reprint requests to: Daniel Herschlag, Department of Biochemistry, Stanford University, Stanford, CA 94305, USA; e-mail: herschla{at}stanford.edu; fax: (650) 723-6783; or Sebastian Doniach, Department of Physics, Stanford University, Stanford, CA 94305; e-mail: doniach{at}drizzle.stanford.edu; fax: (650) 725-2189.
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Article published online ahead of print. Article and publication date are at http://www.rnajournal.org/cgi/doi/10.1261/rna.1985110.
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- Received November 3, 2009.
- Accepted December 4, 2009.
- Copyright © 2010 RNA Society