Structural Insights into Mechanisms of Catalysis and Inhibition in Norwalk Virus Polymerase*

  • Dmitry F. Zamyatkin
    Affiliations
    Department of Biological Sciences and Alberta Ingenuity Centre for Carbohydrate Science, University of Calgary, Calgary, Alberta T2N 1N4, Canada
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  • Francisco Parra
    Affiliations
    Departamento de Bioquímica y Biología Molecular, Instituto Universitario de Biotecnología de Asturias, Universidad de Oviedo, 33006 Oviedo, Spain
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  • José M. Martín Alonso
    Footnotes
    Affiliations
    Departamento de Bioquímica y Biología Molecular, Instituto Universitario de Biotecnología de Asturias, Universidad de Oviedo, 33006 Oviedo, Spain
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  • Daniel A. Harki
    Footnotes
    Affiliations
    Department of Chemistry, The Pennsylvania State University, University Park, Pennsylvania, 16802
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  • Blake R. Peterson
    Affiliations
    Department of Chemistry, The Pennsylvania State University, University Park, Pennsylvania, 16802
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  • Pawel Grochulski
    Affiliations
    Canadian Light Source, Inc., University of Saskatchewan, Saskatoon, Saskatchewan S7N 0X4, Canada
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  • Kenneth K.-S. Ng
    Correspondence
    To whom correspondence should be addressed: Dept. of Biological Sciences, University of Calgary, 2500 University Dr. N.W., Calgary, Alberta T2N 1N4, Canada. Tel.: 403-220-4320; Fax: 403-289-9311
    Affiliations
    Department of Biological Sciences and Alberta Ingenuity Centre for Carbohydrate Science, University of Calgary, Calgary, Alberta T2N 1N4, Canada
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  • Author Footnotes
    * This work was supported in part by Natural Sciences and Engineering Research Council (NSERC), National Research Council, Canadian Institutes of Health Research, the University of Saskatchewan, the Alberta Synchrotron Institute, the National Institutes of Health, the National Science Foundation, the University of California and Henry Wheeler, the Alberta Science and Research Authority, and Alberta Heritage Foundation for Medical Research (AHFMR). This work was supported by NSERC Discovery Grant 262089 and Canadian Institutes of Health Research Operating Grant MOP-67209 (to K. N.), National Institutes of Health Grant AI054776 (to B. R. P.), and Grant BIO2006-00827 from the Spanish Ministerio de Educación y Ciencia cofinanced by Fondo Europeo de Desarrollo Regional (to F. P.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
    1 Recipient of a “Ramón y Cajal” contract from the Spanish Ministerio de Educación y Ciencia cofinanced by Fondo Social Europeo.
    2 Supported by a predoctoral fellowship from the American Heart Association.
      Crystal structures of Norwalk virus polymerase bound to an RNA primer-template duplex and either the natural substrate CTP or the inhibitor 5-nitrocytidine triphosphate have been determined to 1.8Å resolution. These structures reveal a closed conformation of the polymerase that differs significantly from previously determined open structures of calicivirus and picornavirus polymerases. These closed complexes are trapped immediately prior to the nucleotidyl transfer reaction, with the triphosphate group of the nucleotide bound to two manganese ions at the active site, poised for reaction to the 3′-hydroxyl group of the RNA primer. The positioning of the 5-nitrocytidine triphosphate nitro group between the α-phosphate and the 3′-hydroxyl group of the primer suggests a novel, general approach for the design of antiviral compounds mimicking natural nucleosides and nucleotides.

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