1887

Abstract

SUMMARY

The nucleocapsid protein of bovine rotavirus was shown to exist in trimeric units in both the virus particle and in infected cells, with the subunits linked by non-covalent interactions. These trimeric units complex further by disulphide bridges into larger units which may represent the hexameric structures observed by electron microscopy. Visualization of various nucleocapsid protein complexes was also achieved on polyacrylamide gels by treating virus preparations with urea at 37 °C or boiling in the presence and absence of 2-mercaptoethanol. Since virus particles devoid of nucleic acid were also broken down into trimeric subunits by such treatments, assembly of virus particles appears not to require an RNA-protein interaction. Four nucleocapsid-specific monoclonal antibodies with low neutralizing ability reacted with the monomeric (45000 mol. wt., 45K), dimeric (90K), trimeric (135K) and trimeric pair (270K) subunits, indicating that a site responsible for neutralization is probably exposed after assembly of these subunits. Analysis of radiolabelled virus revealed that a high proportion (80 %) of infectious particles could be immunqprecipitated by these monoclonal antibodies, suggesting that the virus particles are either partially doubleshelled or have the nucleocapsid exposed on the surface. The monoclonal antibodies also cross-reacted with the nucleocapsid proteins of simian (SA11), pig (OSU), bovine (NCDV and UK) and human (Wa and ST4) rotaviruses in an immunoblot ELISA reaction. Since these six viruses belong to two different subgroups, it is likely that the antibodies did not recognize the subgroup-specific site, but a shared exposed antigenic determinant. Due to the hexameric configuration of the nucleocapsid in virus particles the neutralizing epitope may be repeatedly presented and, therefore, may contribute to the immunogenicity of this protein.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-68-1-123
1987-01-01
2024-12-06
Loading full text...

Full text loading...

/deliver/fulltext/jgv/68/1/JV0680010123.html?itemId=/content/journal/jgv/10.1099/0022-1317-68-1-123&mimeType=html&fmt=ahah

References

  1. Babiuk L. A., Mohammed K., Spence L., Fauvel M., Petro R. 1977; Rotavirus isolation and cultivation in the presence of trypsin. Journal of Clinical Microbiology 6:610–617
    [Google Scholar]
  2. Bastardo J. w., Mckimm-Breschkin J. L., Sonza S., Mercer L. D., Holmes I. H. 1981; Preparation and characterization of antisera to electrophoretically purified SA11 virus polypeptides. Infection and Immunity 34:641–647
    [Google Scholar]
  3. Bican P., Cohen J., Charp1Llienne A., Scherrer R. 1982; Purification and characterization of bovine rotavirus cores. Journal of Virology 43:1113–1117
    [Google Scholar]
  4. Both G. W., Siegman L. J., Bellamy A. R., Ikegamt N., Shatkin A. J., Furuichi Y. 1984; Comparative sequence analysis of rotavirus genomic segment 6 - the gene specifying viral subgroups 1 and 2. Journal of Virology 51:97–101
    [Google Scholar]
  5. Braun D. K., Pereira L., Norrild B., Roizman B. 1983; Application of denatured, electrophoretically separated, and immobilized lysates of herpes simplex virus-infected cells for detection of monoclonal antibodies and for studies of the properties of viral proteins. Journal of Virology 46:103–112
    [Google Scholar]
  6. Chasey D., Labram J. 1983; Electron microscopy of tubular assemblies associated with naturally occurring bovine rotavirus. Journal of General Virology 64:863–872
    [Google Scholar]
  7. Cleveland D. W., Fischer S. G., Kirschner M. W., Laemmli U. K. 1977; Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. Journal of Biological Chemistry 252:1102–1106
    [Google Scholar]
  8. Cohen J., Laporte J., Charpilienne A., Scherrer R. 1979; Activation of rotavirus RNA polymerase by calcium chelation. Archives of Virology 60:177–186
    [Google Scholar]
  9. Cohen J., Lefevre F., Estes M. K., Bremont M. 1984; Cloning of bovine rotavirus (RF strain): nucleotide sequence of the gene coding for the major capsid protein. Virology 138:178–182
    [Google Scholar]
  10. Estes M. K., Mason B. B., Crawford S., Cohen J. 1984; Cloning and nucleotide sequence of the simian rotavirus gene 6 that codes for the major inner capsid protein. Nucleic Acids Research 12:1875–1887
    [Google Scholar]
  11. Gorziglia M., Larrea C., Liprandi F., Esparza J. 1985; Biochemical evidence for the oligomeric (possibly trimeric) structure of the major inner capsid polypeptide (45K) of rotaviruses. Journal of General Virology 66:1889–1900
    [Google Scholar]
  12. Greenberg H. B., Mcauliffe V., Valdesuso J., Wyatt R. G., Flores J., Kalica A. R., Hoshino Y., Singh N. 1983a; Serological analysis of the subgroup protein of rotavirus, using monoclonal antibodies. Infection and Immunity 39:91–99
    [Google Scholar]
  13. Greenberg H.B, Valdesuso J., Van Wyke K., Midthun K., Walsh M., Mcauliffe V., Wyatt R. G., Kalica A. R., Flores J., Hoshino Y. 1983b; Production and preliminary characterization of monoclonal antibodies directed at two surface proteins of rhesus rotavirus. Journal of Virology 47:267–275
    [Google Scholar]
  14. Hoshino Y., Sereno M. M., Midthun K., Flores J., Kapikian A. Z. 1985; Independent segregation of two antigenic specificities (VP3 and VP7) involved in neutralization of rotavirus infectivity. Proceedings of the National Academy of Sciences, U.S.A 82:8701–8704
    [Google Scholar]
  15. Kalica A. R., Greenberg H. B., Wyatt R. G., Flores J., Sereno M. M., Kapikian A. Z., Chanock R. M. 1981; Genes of human (strain Wa) and bovine (strain UK) rotavirus that code for neutralization and subgroup antigens. Virology 112:385–390
    [Google Scholar]
  16. Kapikian A. Z., Cline W. L., Greenberg H. B., Wyatt R. G., Kalica A. R., Barks C. D., Flores J., Chanock R. M. 1981; Antigenic characterization of human and animal rotavirus by immune adherence hemagglutination assay (I AH A): evidence for distinction of IAH A and neutralization antigens. Infection and Immunity 33:415–425
    [Google Scholar]
  17. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature; London: 227680–685
    [Google Scholar]
  18. Lee P. W. K., Hayes E. C., Joklik W. K. 1981; Protein σ 1 is the reovirus cell attachment protein. Virology 108:156–163
    [Google Scholar]
  19. Martin M. L., Palmer E. L., Middleton P. J. 1975; Ultrastructure of infantile gastroenteritis virus. Virology 68:146–153
    [Google Scholar]
  20. Novo E., Esparza J. 1981; Composition and topography of structural polypeptides of bovine rotavirus. Journal of General Virology 56:325–335
    [Google Scholar]
  21. Offit P. A., Blavat G. 1986; Identification of the two rotavirus genes determining neutralization specificities. Journal of Virology 57:376–378
    [Google Scholar]
  22. Palmer E., Martin M. L. 1982; Further observations on the ultrastructure of human rotavirus. Journal of General Virology 62:105–111
    [Google Scholar]
  23. Palmer E. L, Martin M. L, Murphy F. A. 1977; Morphology and stability of infantile gastroenteritis virus: comparison with reovirus and bluetongue virus. Journal of General Virology 35:403–414
    [Google Scholar]
  24. Petrie B. L., Graham D. Y., Hanssen H., Estes M. K. 1982; Localization of rotavirus antigens in infected cells by ultrastructural immunocytochemistry. Journal of General Virology 63:457–467
    [Google Scholar]
  25. Sabara M., Gilchrist J. E., Hudson G. R., Babiuk L. A. 1985; Preliminary characterization of an epitope involved in neutralization and cell attachment that is located on the major bovine rotavirus glycoprotein. Journal of Virology 53:58–66
    [Google Scholar]
/content/journal/jgv/10.1099/0022-1317-68-1-123
Loading
/content/journal/jgv/10.1099/0022-1317-68-1-123
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