1887

Abstract

A rapid and simple two-step scheme for the purification of herpes simplex virus type 1 UL8 protein from insect cells infected with a recombinant baculovirus has been developed. The scheme involves DEAE–Sepharose and phenyl–Sepharose chromatography and yields approximately 1∙5 mg of protein from 2∙4 × 10 infected cells. The protein remains intact during purification as judged by its reactivity with amino and carboxy termini-specific antisera. Gel filtration chromatography showed that the protein exists as a monomer in solution. No binding of the protein to ssDNA or dsDNA or to a DNA/RNA hybrid could be demonstrated using a gel mobility shift assay.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-74-4-607
1993-04-01
2024-04-25
Loading full text...

Full text loading...

/deliver/fulltext/jgv/74/4/JV0740040607.html?itemId=/content/journal/jgv/10.1099/0022-1317-74-4-607&mimeType=html&fmt=ahah

References

  1. Alberts B. M., Frey L. 1970; T4 bacteriophage gene 32: a structural protein in the replication and recombination of DNA. Nature, London 227:1313–1318
    [Google Scholar]
  2. Alberts B. M., Amodio F., Jenkins M., Guttman E. D., Ferris F. L. 1969; Studies with DNA-cellulose chromatography. I. DNA-binding proteins from Escherichia coli. Cold Spring Harbor Symposia on Quantitative Biology 33:289–305
    [Google Scholar]
  3. Bayliss G. J., Marsden H. S., Hay J. 1975; Herpes simplex virus proteins, DNA-binding proteins in infected cells and in the virus structure. Virology 68:124–134
    [Google Scholar]
  4. Brown M., Faulkner P. 1977; A plaque assay for nuclear polyhedrosis viruses using a solid overlay. Journal of General Virology 36:361–364
    [Google Scholar]
  5. Calder j. M, Stow N. D. 1990; Herpes simplex virus helicase–primase: the UL8 protein is not required for DNA-dependent ATPase and DNA helicase activities. Nucleic Acids Research 36:361–364
    [Google Scholar]
  6. Calder j. M, Stow E. C., Stow N. D. 1992; On the cellular localization of the components of the herpes simplex virus type 1 helicase–primase complex and the viral origin-binding protein. Journal of General Virology 73:531–538
    [Google Scholar]
  7. Crute J. J., Tsurumi T., Zhu L., Weller S. K., Olivo P. D., Challberg M. D., Mocarski E. S., Lehman I. R. 1989; Herpes simplex virus 1 helicase–primase: a complex of three herpes-encoded gene products. Proceedings of the National Academy of Sciences, U.S.A. 86:2186–2189
    [Google Scholar]
  8. Dodson M. S., Lehman I. R. 1991; Association of DNA helicase and primase activities with a subassembly of the herpes simplex virus 1 helicase–primase composed of the UL5 and UL52 gene products. Proceedings of the National Academy of Sciences, U.S.A. 88:1105–1109
    [Google Scholar]
  9. Dorsky D. I., Chatis P., Crumpacker C. 1987; Functional expression of cloned herpes simplex virus type 1 DNA polymerase gene. Journal of Virology 61:1704–1707
    [Google Scholar]
  10. Gallo M. L., Jackwood D. H., Murphy M., Marsden H. S., Parris D. S. 1988; Purification of the herpes simplex virus type 1 65-kilodalton DNA-binding protein: properties of the protein and evidence of its association with the virus-encoded DNA polymerase. Journal of Virology 62:2874–2883
    [Google Scholar]
  11. Gallo M. L., Dorsky D. I., Crumpacker C., Parris D. S. 1989; The essential 65-kilodalton DNA-binding protein of herpes simplex virus stimulates the virus-encoded DNA polymerase. Journal of Virology 63:5023–5029
    [Google Scholar]
  12. Gottlieb J., Marcy A. I., Coen D. M., Challberg M. D. 1990; The herpes simplex virus type 1 UL42 gene product: a subunit of DNA polymerase that functions to increase processivity. Journal of Virology 64:5976–5987
    [Google Scholar]
  13. Hernandez T. R., Lehman I. R. 1990; Functional interaction between the herpes simplex-1 DNA polymerase and UL42 protein. Journal of Biological Chemistry 265:11227–11232
    [Google Scholar]
  14. Kitts P. A., Ayres M. D., Possee R. D. 1990; Linearization of baculovirus DNA enhances the recovery of recombinant vims expression vectors. Nucleic Acids Research 18:5667–5672
    [Google Scholar]
  15. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, London 227:680–685
    [Google Scholar]
  16. McGeoch D. J., Dalrymple M. A., Davison A. J., Dolan A., Frame M. C., McNab D., Perry L. J., Scott J. E., Taylor P. 1988; The complete DNA sequence of the long unique region in the genome of herpes simplex virus type 1. Journal of General Virology 69:1531–1574
    [Google Scholar]
  17. McLean G. W., Owsianka A. M., Subak-Sharpe J. H., Marsden H. S. 1991; Generation of anti-peptide and anti-protein sera: effect of peptide presentation on immunogenicity. Journal of Immunological Methods 137:149–157
    [Google Scholar]
  18. Maniatis T., Fritsch E. F., Sambrook J., Marsden H. S. 1982 Molecular Cloning: A Laboratory Manual New York: Cold Spring Harbor Laboratory;
    [Google Scholar]
  19. Marcy A. I., Olivo P., Challberg M. D., Coen D. M. 1990; Enzymatic activities of overexpressed herpes simplex virus DNA polymerase from recombinant baculovirus-infected insect cells. Nucleic Acids Research 18:1207–1215
    [Google Scholar]
  20. Matsuura Y., Possee R. D., Overton H. A., Bishop DH. L. 1987; Baculovirus expression vectors: the requirements for high level expression of proteins, including glycoproteins. Journal of General Virology 68:1233–1250
    [Google Scholar]
  21. Mocarski E. S., Roizman B. 1982; Herpesvirus-dependent amplification and inversion of cell-associated viral thymidine kinase gene flanked by viral a sequences and linked to an origin of viral DNA replication. Proceedings of the National Academy of Sciences, U.S.A. 79:5626–5630
    [Google Scholar]
  22. Olivo P. D., Nelson N. J., Challberg M. D. 1989; Herpes simplex virus type 1 gene products required for DNA replication: identification and overexpression. Journal of Virology 63:196–204
    [Google Scholar]
  23. Posnett D. N., McGrath H., Tam J. P. 1988; A novel method for producing anti-peptide antibodies. Journal of Biological Chemistry 263:1719–1725
    [Google Scholar]
  24. Quinn J. P., McGeoch D. J. 1985; DNA sequence of the region in the genome of herpes simplex virus type 1 containing the genes for DNA polymerase and the major DNA binding protein. Nucleic Acids Research 13:8143–8163
    [Google Scholar]
  25. Sherman G., Gottlieb J., Challberg M. D. 1992; The UL8 subunit of the herpes simplex virus helicase–primase complex is required for efficient primer utilization. Journal of Virology 66:4884–4892
    [Google Scholar]
  26. Spaete R. R., Frenkel N. 1982; The herpes simplex virus amplicon: a new eucaryotic defective-virus cloning-amplifying vector. Cell 30:295–304
    [Google Scholar]
  27. Stow N. D., McMonagle E. C. 1983; Characterization of the TRS/1RS origin of DNA replication of herpes simplex virus type 1. Virology 130:427–138
    [Google Scholar]
  28. Tam J. P. 1988; Synthetic peptide vaccine design: synthesis and properties of a high-density multiple antigenic peptide system. Proceedings of the National Academy of Sciences, U.S.A. 85:5409–5413
    [Google Scholar]
  29. Towbin H., Staehelin T., Gordon J. 1979; Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proceedings of the National Academy of Sciences, U.S.A. 76:4350–4354
    [Google Scholar]
  30. Tsurimoto T., Stillman B. 1991; Replication factors required for SV40 DNA replication in vitro. 1. DNA structure-specific recognition of a primer-template junction by eukaryotic DNA polymerases and their accessory proteins. Journal of Biological Chemistry 266:1950–1960
    [Google Scholar]
  31. Wang T. SF. 1991; Eukaryotic DNA polymerase. Annual Review of Biochemistry 60:513–552
    [Google Scholar]
  32. Weir H. M., Stow N. D. 1990; Two binding sites for the herpes simplex virus type 1 UL9 protein are required for efficient activity of the oris replication origin. Journal of General Virology 71:1379–1385
    [Google Scholar]
  33. Weir H. M., Calder J. M., Stow N. D. 1989; Binding of the herpes simplex virus type 1 UL9 gene product to an origin of viral DNA replication. Nucleic Acids Research 17:1409–1425
    [Google Scholar]
  34. Weller S. K. 1991; Genetic analysis of HSV genes required for genome replication. In Herpesvirus Transcription and Its Regulation pp 105–135 Edited by Wagner E. K. Boca Raton: CRC Press;
    [Google Scholar]
  35. Weller S. K., Lee K. J., Sabourin D. J., Schaffer P. A. 1983; Genetic analysis of temperature-sensitive mutants which define the gene for the major herpes simplex virus type 1 DNA-binding protein. Journal of Virology 45:354–366
    [Google Scholar]
  36. Weller S. K., Spadaro A., Schaffer J. E., Murray A. W., Maxam A. M., Schaffer P. A. 1985; Cloning, sequencing and functional analysis of orih, a herpes simplex virus type 1 origin of DNA synthesis. Molecular and Cellular Biology 5:930–942
    [Google Scholar]
  37. Wu C. A., Nelson N. J., McGeoch D. J., Challberg M. D. 1988; dentification of herpes simplex virus type 1 genes required for origin-dependent DNA synthesis. Journal of Virology 62:435–443
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-74-4-607
Loading
/content/journal/jgv/10.1099/0022-1317-74-4-607
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error