EFFECTS OF MUTATIONS IN THE EXO-III MOTIF OF THE HERPES-SIMPLEX VIRUS-DNA POLYMERASE GENE ON ENZYME-ACTIVITIES, VIRAL REPLICATION, AND REPLICATION FIDELITY

The herpes simplex virus DNA polymerase catalytic subunit, which has i ntrinsic polymerase and 3'-5' exonuclease activities, contains sequenc e motifs that are homologous to those important for 3'-5' exonuclease activity in other polymerases. The role of one such motif, Eso III, wa s examined in this study, Mutated polymerases containing either a sing le tyrosine-to-histidine change at residue 577 or this change plus an aspartic acid-to-alanine at residue 581 in the fro III motif exhibited defective or undetectable exonuclease activity, respectively, yet ret ained substantial polymerase activity, Despite the defects in exonucle ase activity, the mutant polymerases were able to support viral replic ation in transient complementation assays, albeit inefficiently, Virus es replicated via the action of these mutant polymerases exhibited sub stantially increased frequencies of mutants resistant to ganciclovir, Furthermore, when the fro In mutations were incorporated into the vira l genome, the resulting mutant viruses displayed only modestly defect in replication in Vero cells and exhibited substantially increased mut ation frequencies, The results suggest that herpes simplex virus can r eplicate despite severely impaired exonuclease activity and that the 3 '-5' exonuclease contributes substantially to the fidelity of viral DN A replication.