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RNA-Dependent RNA Polymerase Speed and Fidelity are not the Only Determinants of the Mechanism or Efficiency of Recombination

Department of Biochemistry and Molecular Biology, The Pennsylvania State University, 201 Althouse Laboratory, University Park, PA 16802, USA
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Author to whom correspondence should be addressed.
Present address: Department of Microbiology and Immunology, School of Medicine, University of North Carolina at Chapel Hill, 125 Mason Farm Rd., Chapel Hill, NC 27599-7290, USA.
Genes 2019, 10(12), 968; https://doi.org/10.3390/genes10120968
Received: 15 October 2019 / Revised: 14 November 2019 / Accepted: 21 November 2019 / Published: 25 November 2019
(This article belongs to the Special Issue Virus Evolution and Mutagenesis)
Using the RNA-dependent RNA polymerase (RdRp) from poliovirus (PV) as our model system, we have shown that Lys-359 in motif-D functions as a general acid in the mechanism of nucleotidyl transfer. A K359H (KH) RdRp derivative is slow and faithful relative to wild-type enzyme. In the context of the KH virus, RdRp-coding sequence evolves, selecting for the following substitutions: I331F (IF, motif-C) and P356S (PS, motif-D). We have evaluated IF-KH, PS-KH, and IF-PS-KH viruses and enzymes. The speed and fidelity of each double mutant are equivalent. Each exhibits a unique recombination phenotype, with IF-KH being competent for copy-choice recombination and PS-KH being competent for forced-copy-choice recombination. Although the IF-PS-KH RdRp exhibits biochemical properties within twofold of wild type, the virus is impaired substantially for recombination in cells. We conclude that there are biochemical properties of the RdRp in addition to speed and fidelity that determine the mechanism and efficiency of recombination. The interwoven nature of speed, fidelity, the undefined property suggested here, and recombination makes it impossible to attribute a single property of the RdRp to fitness. However, the derivatives described here may permit elucidation of the importance of recombination on the fitness of the viral population in a background of constant polymerase speed and fidelity. View Full-Text
Keywords: enterovirus; RNA-dependent RNA polymerase (RdRp); nucleotidyl transfer mechanism; general acid; fidelity; recombination; copy-choice; forced-copy-choice enterovirus; RNA-dependent RNA polymerase (RdRp); nucleotidyl transfer mechanism; general acid; fidelity; recombination; copy-choice; forced-copy-choice
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Kim, H.; Ellis, V.D., III; Woodman, A.; Zhao, Y.; Arnold, J.J.; Cameron, C.E. RNA-Dependent RNA Polymerase Speed and Fidelity are not the Only Determinants of the Mechanism or Efficiency of Recombination. Genes 2019, 10, 968.

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