title: UL52 primase interactions in the herpes simplex virus 1 helicase-primase are affected by antiviral compounds and mutations causing drug resistance.
abstract: Herpes simplex virus 1 (HSV-1) UL5/8/52 helicase-primase complex is required for DNA unwinding at the replication fork and synthesis of primers during virus replication, and it has become a promising novel target for antiviral therapy. Using molecular cloning, we have identified three separate domains of UL52. Co-immunoprecipitation experiments in extracts from cells transiently expressing HA-tagged UL5, FLAG-UL8, and enhanced GFP-tagged UL52 domains revealed that the N-terminal domain of UL52 primase binds UL5 helicase and the middle domain interacts with the UL8 accessory protein. In addition, an interaction between the single strand DNA-binding protein ICP8 and the UL52 middle domain was observed. The complex between UL5 and UL52 was stabilized by the antiviral compound BAY 54-6322, and mutations providing resistance to the drug obliterate this effect. Our results also suggest a mechanism for accommodating conformational strain resulting from movement of UL5 and UL52 in opposite directions on the lagging strand template, and they identify molecular complexes that can be further examined by structural biology techniques to resolve the mechanism of primer synthesis during herpesvirus replication. Finally, they help to explain the mechanism of action of a novel class of antiviral compounds currently being evaluated in clinical trials.
authors.affiliation: From the Institute of Biomedicine, Department of Medical Biochemistry and Cell Biology, Sahlgrenska Academy, University of Gothenburg, Box 440, SE-405 30 Gothenburg, Sweden.
authors.forename: Isabella
authors.initials: I
authors.lastname: Muylaert
authors.affiliation: From the Institute of Biomedicine, Department of Medical Biochemistry and Cell Biology, Sahlgrenska Academy, University of Gothenburg, Box 440, SE-405 30 Gothenburg, Sweden.
authors.forename: Zhiyuan
authors.initials: Z
authors.lastname: Zhao
authors.affiliation: From the Institute of Biomedicine, Department of Medical Biochemistry and Cell Biology, Sahlgrenska Academy, University of Gothenburg, Box 440, SE-405 30 Gothenburg, Sweden per.elias@medkem.gu.se.
authors.forename: Per
authors.initials: P
authors.lastname: Elias
mesh_terms: D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D002460:Cell Line; D004265:DNA Helicases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D019915:DNA Primase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication / Q000187:drug effects / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D024882:Drug Resistance, Viral / Q000187:drug effects* / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D018259:Herpesvirus 1, Human / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D018613:Microscopy, Confocal; D009154:Mutation; D010069:Oxadiazoles / Q000494:pharmacology; D011485:Protein Binding; D014764:Viral Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication / Q000187:drug effects / Q000235:genetics
chemical_list: C568714:ASP2151; D000998:Antiviral Agents; D004268:DNA-Binding Proteins; C045462:ICP8 protein, Simplexvirus; D010069:Oxadiazoles; D014764:Viral Proteins; D049452:Green Fluorescent Proteins; D019915:DNA Primase; C062021:helicase-primase, Human herpesvirus 1; D004265:DNA Helicases
keywords: Antiviral Agent; DNA Primase; DNA Replication; Herpesvirus; Viral Replication