title: A point mutation within conserved region VI of herpes simplex virus type 1 DNA polymerase confers altered drug sensitivity and enhances replication fidelity.
abstract: Herpes simplex virus type 1 (HSV-1) DNA polymerase contains several conserved regions within the polymerase domain. The conserved regions I, II, III, V, and VII have been shown to have functional roles in the interaction with deoxynucleoside triphosphates (dNTPs) and DNA. However, the role of conserved region VI in DNA replication has remained unclear due, in part, to the lack of a well-characterized region VI mutant. In this report, recombinant viruses containing a point mutation (L774F) within the conserved region VI were constructed. These recombinant viruses were more susceptible to aphidicolin and resistant to both foscarnet and acyclovir, compared to the wild-type KOS strain. Marker transfer experiments demonstrated that the L774F mutation conferred the altered drug sensitivities. Furthermore, mutagenesis assays demonstrated that L774F recombinant viruses containing the supF marker gene, which was integrated within the thymidine kinase locus (tk), exhibited increased fidelity of DNA replication. These data indicate that conserved region VI, together with other conserved regions, forms the polymerase active site, has a role in the interaction with deoxyribonucleotides, and regulates DNA replication fidelity. The possible effect of the L774F mutation in altering the polymerase structure and activity is discussed.
authors.affiliation: Department of Microbiology and Immunology, College of Medicine, SUNY, Upstate Medical University, Syracuse, New York 13210, USA.
authors.forename: Ying T
authors.initials: YT
authors.lastname: Hwang
authors.forename: Harmon J
authors.initials: HJ
authors.lastname: Zuccola
authors.forename: Qiaosheng
authors.initials: Q
authors.lastname: Lu
authors.forename: Charles B C
authors.initials: CB
authors.lastname: Hwang
mesh_terms: D000212:Acyclovir / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D016590:Aphidicolin / Q000494:pharmacology; D002460:Cell Line; D002522:Chlorocebus aethiops; D017124:Conserved Sequence; D004261:DNA Replication; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D024882:Drug Resistance, Viral; D017245:Foscarnet / Q000494:pharmacology; D018259:Herpesvirus 1, Human / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D017354:Point Mutation; D016415:Sequence Alignment; D014709:Vero Cells; D010948:Viral Plaque Assay
chemical_list: D000998:Antiviral Agents; D017245:Foscarnet; D016590:Aphidicolin; D004259:DNA-Directed DNA Polymerase; D000212:Acyclovir