Expression of the varicella-zoster virus origin-binding protein and analysis of its site-specific DNA-binding properties.
The varicella-zoster virus (VZV) genome contains homologs to each of the seven herpes simplex virus (HSV) genes that are required for viral DNA synthesis. VZV gene 51 is homologous to HSV UL9, which encodes an origin of DNA replication binding protein (OBP). It was previously shown, by using a protein A fusion protein, that the product of gene 51 is a site-specific DNA-binding protein which binds to sequences within the VZV origin (Stow et al., Virology 177:570-577, 1990). In this report, gene 51 was expressed in an in vitro translation system. Rabbit antiserum raised against the carboxyl-terminal 20 amino acids was used to confirm expression of the full-length gene 51 protein, and site-specific DNA-binding activity was demonstrated in a gel retardation assay. The origin-binding domain was located within a 263-amino-acid region of the carboxyl terminus by using a series of deletion mutants. The affinity of binding of the VZV OBP to the three binding sites in the VZV origin was found to be similar. In addition, as with UL9, a CGC triplet within a 10-bp consensus sequence is critical to the interaction between the OBP and the origin. The HSV and VZV OBPs, therefore, appear to have virtually identical recognition sequences despite only 33% identity and 44% similarity in the primary structure of their site-specific DNA-binding domains.
Chen
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Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
Olivo
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D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral; D001483:Base Sequence; D001665:Binding Sites; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015870:Gene Expression; D005814:Genes, Viral; D014645:Herpesvirus 3, Human / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015345:Oligonucleotide Probes / Q000235:genetics; D017354:Point Mutation; D014176:Protein Biosynthesis; D011817:Rabbits; D017386:Sequence Homology, Amino Acid; D018139:Simplexvirus / Q000235:genetics; D013045:Species Specificity; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
D000914:Antibodies, Viral; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D015345:Oligonucleotide Probes; D014764:Viral Proteins; C055789:origin-binding proteins, viral