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

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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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The atomic coordinates and structure factors (code 3BSN and 3BSO) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

*

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.