title: Remote site control of an active site fidelity checkpoint in a viral RNA-dependent RNA polymerase.
abstract: The kinetic, thermodynamic, and structural basis for fidelity of nucleic acid polymerases remains controversial. An understanding of viral RNA-dependent RNA polymerase (RdRp) fidelity has become a topic of considerable interest as a result of recent experiments that show that a 2-fold increase in fidelity attenuates viral pathogenesis and a 2-fold decrease in fidelity reduces viral fitness. Here we show that a conformational change step preceding phosphoryl transfer is a key fidelity checkpoint for the poliovirus RdRp (3Dpol). We provide evidence that this conformational change step is orientation of the triphosphate into a conformation suitable for catalysis, suggesting a kinetic and structural model for RdRp fidelity that can be extrapolated to other classes of nucleic acid polymerases. Finally, we show that a site remote from the catalytic center can control this checkpoint, which occurs at the active site. Importantly, similar connections between a remote site and the active site exist in a wide variety of viral RdRps. The capacity for sites remote from the catalytic center to alter fidelity suggests new possibilities for targeting the viral RdRp for antiviral drug development.
authors.affiliation: Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA.
authors.forename: Jamie J
authors.initials: JJ
authors.lastname: Arnold
authors.forename: Marco
authors.initials: M
authors.lastname: Vignuzzi
authors.forename: Jeffrey K
authors.initials: JK
authors.lastname: Stone
authors.forename: Raul
authors.initials: R
authors.lastname: Andino
authors.forename: Craig E
authors.initials: CE
authors.lastname: Cameron
mesh_terms: D000249:Adenosine Monophosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites / Q000502:physiology; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004789:Enzyme Activation / Q000235:genetics; D014644:Genetic Variation; D006367:HeLa Cells; D006801:Humans; D009711:Nucleotides / Q000378:metabolism; D010766:Phosphorylation; D017354:Point Mutation; D017955:Poliovirus / Q000201:enzymology* / Q000235:genetics*; D017434:Protein Structure, Tertiary; D012367:RNA, Viral / Q000502:physiology*; D013329:Structure-Activity Relationship
chemical_list: D009711:Nucleotides; D012367:RNA, Viral; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D012321:DNA-Directed RNA Polymerases