Aphidicolin resistance in herpes simplex virus type I reveals features of the DNA polymerase dNTP binding site.
We describe the mapping and sequencing of mutations within the DNA polymerase gene of herpes simplex virus type 1 which confer resistance to aphidicolin, a DNA polymerase inhibitor. The mutations occur near two regions which are highly conserved among DNA polymerases related to the herpes simplex enzyme. They also occur near other herpes simplex mutations which affect the interactions between the polymerase and deoxyribonucleoside triphosphate substrates. Consequently, we argue in favor of the idea that the aphidicolin binding site overlaps the substrate binding site and that the near-by conserved regions are functionally required for substrate binding. Our mutants also exhibit abnormal sensitivity to another DNA polymerase inhibitor, phosphonoacetic acid. This drug is thought to bind as an analogue of pyrophosphate. A second-site mutation which suppresses the hypersensitivity of one mutant to phosphonoacetic acid (but not its aphidicolin resistance) is described. This second mutation may represent a new class of mutations, which specifically affects pyrophosphate, but not substrate, binding.
Hall
J D
JD
Department of Molecular and Cellular Biology, University of Arizona, Tucson 85721.
Wang
Y S
YS
Pierpont
J
J
Berlin
M S
MS
Rundlett
S E
SE
Woodward
S
S
D000595:Amino Acid Sequence; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D016590:Aphidicolin; D001483:Base Sequence; D001665:Binding Sites; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D003854:Deoxyribonucleotides / Q000378:metabolism; D004224:Diterpenes / Q000494:pharmacology*; D004352:Drug Resistance, Microbial; D005814:Genes, Viral; D008969:Molecular Sequence Data; D009154:Mutation; D010746:Phosphonoacetic Acid / Q000494:pharmacology; D015183:Restriction Mapping; D018139:Simplexvirus / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D014709:Vero Cells; D010948:Viral Plaque Assay; D015678:Viral Structural Proteins / Q000235:genetics*
D000998:Antiviral Agents; D003854:Deoxyribonucleotides; D004224:Diterpenes; D015678:Viral Structural Proteins; D016590:Aphidicolin; D004259:DNA-Directed DNA Polymerase; D010746:Phosphonoacetic Acid