1887

Abstract

Recombinant baculoviruses have been constructed which express the hepatitis C virus (HCV) NS3 proteinase and its substrates in insect cells. The expressed proteinase has been shown to carry out -cleavage at the NS3/4A, NS4A/4B, NS4B/5A and NS5A/5B junctions in a cell-based assay. When assayed in a cell-free system using translated substrates, the proteinase could perform -processing of the NS4A/4B and NS5A/5B junctions, but only when coexpressed with NS4A, either as an NS3-4A precursor or by co-infection of cells with NS3- and NS4A-expressing recombinant baculoviruses. Possible reasons for the absolute requirement of the NS3 proteinase for NS4A are discussed.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-76-12-3009
1995-12-01
2021-10-16
Loading full text...

Full text loading...

/deliver/fulltext/jgv/76/12/JV0760123009.html?itemId=/content/journal/jgv/10.1099/0022-1317-76-12-3009&mimeType=html&fmt=ahah

References

  1. Bartenschlager R., Ahlborn-Laake L., Mous J., Jacobsen H. 1993; Nonstructural protein 3 of the hepatitis C virus encodes aserine proteinase required for cleavage at the NS3/4 and NS4/5junctions. Journal of Virology 67:3835–3844
    [Google Scholar]
  2. Bartenschlager R., Ahlborn-Laake L., Mous J., Jacobsen H. 1994; Kinetic and structural analyses of hepatitis C virus polyprotein processing. Journal of Virology 68:5045–5055
    [Google Scholar]
  3. Bouffard P., Bartenschlager R., Ahlborn-Laake L., Mous J., Roberts N., Jacobsen H. 1995; An in vitro assay for hepatitis Cvirus proteinase. Virology (in press)
    [Google Scholar]
  4. Brown H., Faulkner P. 1977; A plaque assay for nuclear polyhedrosis viruses using a solid overlay. Journal of General Virology 36:361–364
    [Google Scholar]
  5. Chambers T. J., Weir R. C., Grakoui A., McCourt D. W., Bazan J. F., Fletterick R. J., Rice C. M. 1990; Evidence that the N-terminal domain of nonstructural protein NS3 from yellow fever virus is a serine proteinase responsible for site-specific cleavages in the viral polyprotein. Proceedings of the National Academy of Sciences, USA 87:8898–8902
    [Google Scholar]
  6. Choo Q.-L., Richman K. H., Han J. H., Berger K., Lee C., Dong C., Gallegos C., Coit D., Medina-Selby A., Barr P. J., Weiner A. J., Bradley D. W., Kuo G., Houghton M. 1991; Genetic organization and diversity of the hepatitis C virus. Proceedings of the National Academy of Sciences, USA 88:2451–2455
    [Google Scholar]
  7. Eckart M. R., Selby M., Masiarz F., Lee C., Berger K., Crawford K., Kuo C., Kuo G., Houghton M., Choo Q.-L. 1993; The hepatitis C virus encodes a serine proteinase involved in processing of the putative nonstructural proteins from the viral polyprotein precursor. Biochemical and Biophysical Research Communications 192:399–406
    [Google Scholar]
  8. Failla C., Tomei L., De Francesco R. 1994; Both NS3 andNS4A are required for proteolytic processing of hepatitis C virus nonstructural proteins. Journal of Virology 68:3753–3760
    [Google Scholar]
  9. Failla C., Tomei L., De Francesco R. 1995; An amino-terminal domain of the hepatitis C virus NS3 protease is essential for interaction with NS4A. Journal of Virology 69:1769–1777
    [Google Scholar]
  10. Fung M.-C., Chiu K. Y. M., Weber T., Chang T. -W., Chang N. T. 1988; Detection and purification of a recombinant human Blymphotropic virus (HHV-6) in the baculovirus expression system by limiting dilution and DNA dot-blot hybridization. Journal of Virological Methods 19:33–42
    [Google Scholar]
  11. Grakoui A., McCourt D. W., Wychowski C., Feinstone S. M., Rice C. M. 1993; Characterization of the hepatitis C virus-encoded serine proteinase: determination of proteinase-dependent polyprotein cleavage sites. Journal of Virology 67:2832–2843
    [Google Scholar]
  12. Hijikata M., Mizushima H., Tanji Y., Komoda Y., Hirowatari Y., Akagi T., Kato N., Kimura K., Shimotohno K. 1993; Proteolytic processing and membrane association of putative nonstructural proteins of hepatitis C virus. Proceedings of the National Academy of Sciences, USA 90:10773–10777
    [Google Scholar]
  13. Hirowatari Y., Hijikata M., Tanji Y., Nyunoya H., Mizushima H., Kimura K., Tanaka T., Kato N., Shimotohno K. 1993; Two proteinase activities in HCV polypeptide expressed in insect cells using baculovirus vector. Archives of Virology 133:349–356
    [Google Scholar]
  14. Kajo N., Hijikata M., Ootsuyama Y., Nakagawa M., Ohkoshi S., Sugimura T., Shimotohno K. 1990; Molecular cloning of the human hepatitis C virus genome from Japanese patients with non-A, non-B hepatitis. Proceedings of the National Academy of Sciences, USA 87:9524–9528
    [Google Scholar]
  15. Knudson D. L., Tinsley T. W. 1974; Replication of a nuclear polyhedrosis virus in a continuous cell culture of Spodoptera frugiperda: purification, assay of infectivity and growth characteristics of the virus. Journal of Virology 14:934–944
    [Google Scholar]
  16. Lin C., Pragai B., Grakoui A., Xu J., Rice C. M. 1994; Hepatitis C virus NS3 serine proteinase: trans-cleavage requirements and processing kinetics. Journal of Virology 68:8147–815
    [Google Scholar]
  17. Manabe S., Fuke I., Tanashita O., Kaji C., Gomi Y., Yoshida S., Mori C., Takamizawa A., Yosida I., Okayama H. 1994; Production of nonstructural proteins of hepatitis C virus requires aputative viral protease encoded by NS3. Virology 198:636–644
    [Google Scholar]
  18. Matsuura Y., Possee R. D., Overton H. A., Bishop D. H. L. 1987; Baculovirus expression vectors: the requirements for high-level expression of proteins, including glycoproteins. Journal of General Virology 68:1233–1250
    [Google Scholar]
  19. Tanji Y., Hijikata M., Satoh S., Kaneko T., Shimotohno K. 1995; Hepatitis C virus nonstructural protein NS4A has versatilefunctions in viral protein processing. Journal of Virology 69:1575–1581
    [Google Scholar]
  20. Tomei L., Failla C., Santolini E., De Francesco R., Lamonica N. 1993; NS3 is a serine protease required for processing of the patitis C virus poly protein. Journal of Virology 674017–4026
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-76-12-3009
Loading
/content/journal/jgv/10.1099/0022-1317-76-12-3009
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

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