RNA-dependent RNA polymerase speed and fidelity are not the only determinants of the mechanism or efficiency of recombination

Hyejeong Kim; Victor D. Ellis; Andrew Woodman; Yan Zhao; Jamie J. Arnold; Craig E. Cameron

For more than a decade now, our laboratory and others have published studies asserting 234 connections between viral RdRp fidelity and viral fitness (10-15). However, in the few instances in 235 which RdRp derivatives exhibiting perturbed fidelity have been characterized, these derivatives also 236 exhibit changes to the rate of nucleotide addition (6, 10, 17, 22, 33). Relative to wild-type polymerase, 237 a higher fidelity polymerase is a slower enzyme, and a lower fidelity polymerase is a faster enzyme (6, 238 10, 17, 22, 33). Polymerase speed will clearly determine replication kinetics and therefore can 239 contribute to viral fitness, as has been suggested recently (17). The most extensively characterized 240 fidelity mutants of PV harbor an RdRp with a substitution located at a remote site that likely causes 241 substantial collateral damage, further confounding the fidelity-versus-speed debate (4, 16). This study 242 was motivated by the need to understand better the relationship between the biochemical properties of 243 the viral RdRp and viral fitness, virulence, and pathogenesis. We have had in hand for a long time a 244 PV mutant whose speed and fidelity were perturbed by changing an RdRp active-site residue, K359H 245 (24). We did not publish this PV mutant until now because of its genetic instability (Fig. 1A) . We 246 realized that this genetic instability might actually represent an opportunity, as the biochemical 247 phenotypes reverted by the second-site suppressors might highlight the biochemical properties driving 248 viral fitness.