1887

Abstract

The DNA polymerase from nucleopolyhedrovirus (SpliNPV) was expressed in, and purified from, prokaryotic and eukaryotic expression systems. While less protein was obtained from the expression system, SpliNPV DNAPOL purified from displayed similar biochemical characteristics to DNAPOL expressed in, and subsequently purified from, insect cells (Sf9) using a baculovirus expression system. Biochemical analyses suggested that the DNA polymerase and the 3′–5′ exonuclease activities are intrinsic to the protein. Deletion of the first 80 amino acid residues from the N terminus of the DNAPOL affected neither the DNA polymerase nor the exonuclease activities of the enzyme. Replication products from single-stranded M13 DNA demonstrated that the DNA synthesis activity of SpliNPV DNAPOL is highly processive. Transient expression assays with a set of deletion clones containing the putative SpliNPV non- origin of DNA replication permitted functional characterization of sequence elements within the origin fragment. Purified SpliNPV DNAPOL stimulated origin-dependent DNA replication in a cell-free replication assay.

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2001-07-01
2020-07-09
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References

  1. Ahrens C. H., Russell R. L., Funk C. J., Evans J. T., Harwood S. H., Rohrmann G. F.. 1997; The sequence of the Orgyia pseudotsugata multinucleocapsid nuclear polyhedrosis virus genome. Virology229:381–399
    [Google Scholar]
  2. Ayres M. D., Howard S. C., Kuzio J., Lopez-Ferber M., Possee R. D.. 1994; The complete DNA sequence of Autographa californica nuclear polyhedrosis virus. Virology202:586–605
    [Google Scholar]
  3. Bradford M. M.. 1976; Rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry72:248–254
    [Google Scholar]
  4. Bulach D. M., Kumar C. A., Zaia A., Liang B., Tribe D. E.. 1999; Group II nucleopolyhedrovirus subgroups revealed by phylogenetic analysis of polyhedrin and DNA polymerase gene sequences. Journal of Invertebrate Pathology73:59–73
    [Google Scholar]
  5. DePamphilis M. L.. 1996; Eukaryotic replication origins. In DNA Replication in Eukaryotic Cells pp45–86 Edited by DePamphilis M. L.. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  6. Gomi S., Majima K., Maeda S.. 1999; Sequence analysis of the genome of Bombyx mori nucleopolyhedrovirus. Journal of General Virology80:1323–1337
    [Google Scholar]
  7. Habib S., Hasnain S. E.. 2000; Differential activity of two non- hr origins during replication of the baculovirus Autographa californica nuclear polyhedrosis virus genome. Journal of Virology74:5182–5189
    [Google Scholar]
  8. Hang X., Guarino L. A.. 1999; Purification of Autographa californica nucleopolyhedrovirus DNA polymerase from infected insect cells. Journal of General Virology80:2519–2526
    [Google Scholar]
  9. Heldens J. G. M., Broer R., Zuidema D., Goldbach R. W., Vlak J. M.. 1997; Identification and functional analysis of a non- hr origin of DNA replication in the genome of Spodoptera exigua nucleopolyhedrovirus. Journal of General Virology78:1497–1506
    [Google Scholar]
  10. Huang J., Levin D. B.. 1999; Identification and functional analysis of a putative non- hr origin of DNA replication from the Spodoptera littoralis multinucleocapsid nucleopolyhedrovirus. Journal of General Virology80:2263–2274
    [Google Scholar]
  11. Huang J., Levin D. B.. 2001; Identification, transcription and sequence analysis of the Spodoptera littoralis nucleopolyhedrovirus DNA polymerase gene. Archives of Virology146:303–326
    [Google Scholar]
  12. IJkel W. F., van Strien E. A., Heldens J. G., Broer R., Zuidema D., Goldbach R. W., Vlak J. M.. 1999; Sequence and organization of the Spodoptera exigua multicapsid nucleopolyhedrovirus genome. Journal of General Virology80:3289–3304
    [Google Scholar]
  13. Kool M., Ahrens C. H., Goldbach R. W., Rohrmann G. F., Vlak J. M.. 1994; Identification of genes involved in DNA replication of the Autographa californica baculovirus. Proceedings of the National Academy of Sciences, USA91:11212–11216
    [Google Scholar]
  14. Kool M., Ahrens C. H., Vlak J. M., Rohrmann G. F.. 1995; Replication of baculovirus DNA. Journal of General Virology76:2103–2118
    [Google Scholar]
  15. Kuzio J., Pearson M. N., Harwood S. H., Funk C. J., Evans J. T., Slavicek J. M., Rohrmann G. F.. 1999; Sequence and analysis of the genome of a baculovirus pathogenic for Lymantria dispar . Virology253:17–34
    [Google Scholar]
  16. Laemmli U. K.. 1970; Cleavage of structural proteins during the assembly of the head of bacterial phage T4. Nature277:680–688
    [Google Scholar]
  17. Lee H. J., Krell P. J.. 1994; Reiterated DNA fragments in defective genomes of Autographa californica nuclear polyhedrosis virus are competent for AcMNPV dependent DNA replication. Virology202:418–429
    [Google Scholar]
  18. Lu A., Krell P. J., Vlak J. M., Rohrmann G. F.. 1997; Baculovirus DNA replication. In The Baculoviruses pp171–191 Edited by Miller L. K.. New York: Plenum Press;
    [Google Scholar]
  19. McDougal V. V., Guarino L. A.. 1999; Autographa californica nuclear polyhedrosis virus DNA polymerase: measurements of processivity and strand displacement. Journal of Virology73:4908–4918
    [Google Scholar]
  20. Mikhailov V. S., Marlyev K. A., Ataeva J. O., Kullyev P. K., Atrazhev A. M.. 1986; Characterization of 3′ to 5′ exonuclease associated with DNA polymerase of silkworm nuclear polyhedrosis virus. Nucleic Acids Research14:3841–3857
    [Google Scholar]
  21. O’Reilly D. R., Miller L. K., Luckow V. A.. 1992; Baculovirus Expression Vectors. A Laboratory Manual New York: W. H. Freeman;
    [Google Scholar]
  22. Pearson M. N., Bjornson R. M., Ahrens C., Rohrmann G. F.. 1993; Identification and characterization of a putative origin of DNA replication in the genome of a baculovirus pathogenic for Orgyia pseudotsugata . Virology197:715–725
    [Google Scholar]
  23. Pedrali-Noy G., Spadari S.. 1980; Mechanism of inhibition of herpes simplex virus and vaccinia virus DNA polymerases by aphidicolin, a highly specific inhibitor of DNA replication in eucaryotes. Journal of Virology36:457–464
    [Google Scholar]
  24. Sambrook J., Fritsch E. F., Maniatis T.. 1989; Molecular Cloning: A Laboratory Manual , 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  25. Sarisky R. T., Hayward G. S.. 1996; Evidence that the UL84 gene product of human cytomegalovirus is essential for promoting ori Lyt-dependent DNA replication and formation of replication compartments in cotransfection assays. Journal of Virology70:7398–7413
    [Google Scholar]
  26. Schumacher S. B., Stucki M., Hubscher U.. 2000; The N-terminal region of DNA polymerase delta catalytic subunit is necessary for holoenzyme function. Nucleic Acids Research28:620–625
    [Google Scholar]
  27. Sheaff R., Ilsley D., Kuchta R.. 1991; Mechanism of DNA polymerase alpha inhibition by aphidicolin. Biochemistry30:8590–8597
    [Google Scholar]
  28. Stillman B., Gluzman Y.. 1985; Replication and supercoiling of simian virus 40 DNA in cell extracts from human cells. Molecular and Cellular Biology5:2051–2060
    [Google Scholar]
  29. Volkman L. E., Blissard G. W., Freisen P. D., Possee R. D., Theilmann D. A.. 1995; The Baculoviridae . In Virus Taxonomy. Sixth Report of the International Committee on Taxonomy of Viruses pp104–113 Edited by Murphy F. A., Fauquet C. M., Bishop D. H. L., Ghabrial S. A., Jarvis A. W., Martelli G. P., Mayo M. A., Summers M. D. Vienna & New York: Springer-Verlag;
    [Google Scholar]
  30. Zanotto P. M., Kessing B. D., Maruniak J. E.. 1993; Phylogenetic interrelationships among baculoviruses: evolutionary rates and host associations. Journal of Invertebrate Pathology62:147–164
    [Google Scholar]
  31. Zhang S. J., Zeng X. R., Zhang P., Toomey N. L., Chuang R. Y., Chang L. S., Lee M. Y.. 1995; A conserved region in the amino terminus of DNA polymerase delta is involved in proliferating cell nuclear antigen binding. Journal of Biological Chemistry270:7988–7992
    [Google Scholar]
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