Expression, purification, and functional characterization of the DNA-binding domain of the herpes simplex virus type 1 UL9 protein.
The origin-binding protein (UL9 protein) encoded by the herpes simplex virus type 1 (HSV-1) UL9 gene plays a critical role in the initiation of viral DNA replication. The 317-amino acid C-terminal DNA-binding domain (UL9-COOH) of the UL9 protein was expressed as a fusion protein to glutathione S-transferase in bacteria. Milligram quantities of the fusion protein were purified by differential ammonium sulfate precipitation and affinity chromatography over reduced glutathione agarose. The UL9-COOH moiety was cleaved from the fusion protein by proteolysis with thrombin. The purified UL9-COOH displayed specific and stable binding activity toward its DNA recognition sequences within the HSV-1 origin of DNA replication.
Martinez
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Drug Discovery Research Group, Genelabs Incorporated, Redwood City, California 94063.
Edwards
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CA
D001483:Base Sequence; D001665:Binding Sites; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D003851:Deoxyribonucleases / Q000378:metabolism; D004926:Escherichia coli; D005982:Glutathione Transferase / Q000235:genetics; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013917:Thrombin; D014764:Viral Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; C064565:UL9 protein, Human herpesvirus 1; D004247:DNA; D005982:Glutathione Transferase; D003851:Deoxyribonucleases; D013917:Thrombin