A fast selenium derivatization strategy for crystallization and phasing of RNA structures

  1. Vincent Olieric1,5,
  2. Ulrike Rieder2,5,
  3. Kathrin Lang2,
  4. Alexander Serganov3,
  5. Clemens Schulze-Briese1,
  6. Ronald Micura2,
  7. Philippe Dumas4,6 and
  8. Eric Ennifar4,6
  1. 1Swiss Light Source, Paul Scherrer Institute, CH-5232 Villigen, Switzerland
  2. 2Institute of Organic Chemistry, Center for Molecular Biosciences, Leopold Franzens University, A-6020 Innsbruck, Austria
  3. 3Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA
  4. 4Architecture et Réactivité de l'ARN, Institut de Biologie Moléculaire et Cellulaire, Université Louis Pasteur/CNRS UPR 9002, F-67084 Strasbourg, France

    1. ↵5 These authors contributed equally to this work.

    Abstract

    Site-specific 2′-methylseleno RNA labeling is a promising tool for tackling the phase problem in RNA crystallography. We have developed an efficient strategy for crystallization and structure determination of RNA and RNA/protein complexes based on preliminary crystallization screening of 2′-OCH3-modified RNA sequences, prior to the replacement of 2′-OCH3 groups with their 2′-SeCH3 counterparts. The method exploits the similar crystallization properties of 2′-OCH3- and 2′-SeCH3-modified RNAs and has been successfully validated for two test cases. In addition, our data show that 2′-SeCH3-modified RNA have an increased resistance to X-ray radiolysis in comparison with commonly used 5-halogen-modified RNA, which permits collection of experimental electron density maps of remarkable quality.

    Keywords:

    Keywords

    Footnotes

    • ↵6 Present address: Architecture et Réactivité de l'ARN, Université de Strasbourg, CNRS, IBMC, 15 rue René Descartes, F-67084 Strasbourg, France.

    • Reprint requests to: Eric Ennifar, Architecture et Réactivité de l'ARN, Institut de Biologie Moléculaire et Cellulaire, Université Louis Pasteur/CNRS UPR 9002, 15 Rue René Descartes, F-67084 Strasbourg, France; e-mail: e.ennifar{at}ibmc.u-strasbg.fr; fax: 33-3-88602218.

    • Article published online ahead of print. Article and publication date are at http://www.rnajournal.org/cgi/doi/10.1261/rna.1499309.

      • Received December 5, 2008.
      • Accepted January 9, 2009.
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    1. Published in Advance February 18, 2009, doi: 10.1261/rna.1499309 RNA 15: 707-715 Copyright © 2009 RNA Society

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