Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1988 May;56(5):1237–1241. doi: 10.1128/iai.56.5.1237-1241.1988

Heterologous expression of the 65-kilodalton antigen of Mycobacterium leprae and murine T-cell responses to the gene product.

F I Lamb 1, A E Kingston 1, I Estrada 1, M J Colston 1
PMCID: PMC259796  PMID: 3281904

Abstract

The gene encoding the immunodominant 65-kilodalton antigen of Mycobacterium leprae was subcloned from a lambda gt11 clone into the high-copy-number plasmid pUC8. Escherichia coli containing these recombinants produced large amounts of the antigen, which was purified by polyacrylamide gel electrophoresis in the presence of urea. The ability of E. coli to recognize the mycobacterial promoter was confirmed by constructing additional clones in which the gene is flanked by transcriptional terminators from phage fd. A similar approach was used to demonstrate the expression of this gene in Streptomyces lividans. Mice immunized with killed M. leprae showed cell-mediated immune reactivity to the purified 65-kilodalton protein which stimulated both in vitro lymphoproliferative and in vivo delayed-type hypersensitivity responses.

Full text

PDF
1237

Images in this article

1239Image
on p.1239
1239Image
on p.1239

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Cox R. A., Katoch V. M. Evidence for genetic divergence in ribosomal RNA genes in mycobacteria. FEBS Lett. 1986 Jan 20;195(1-2):194–198. doi: 10.1016/0014-5793(86)80159-4. [DOI] [PubMed] [Google Scholar]
  2. Emmrich F., Thole J., van Embden J., Kaufmann S. H. A recombinant 64 kilodalton protein of Mycobacterium bovis bacillus Calmette-Guerin specifically stimulates human T4 clones reactive to mycobacterial antigens. J Exp Med. 1986 Apr 1;163(4):1024–1029. doi: 10.1084/jem.163.4.1024. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Gillis T. P., Buchanan T. M. Production and partial characterization of monoclonal antibodies to Mycobacterium leprae. Infect Immun. 1982 Jul;37(1):172–178. doi: 10.1128/iai.37.1.172-178.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Gillis T. P., Miller R. A., Young D. B., Khanolkar S. R., Buchanan T. M. Immunochemical characterization of a protein associated with Mycobacterium leprae cell wall. Infect Immun. 1985 Aug;49(2):371–377. doi: 10.1128/iai.49.2.371-377.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Kaufmann S. H., Väth U., Thole J. E., Van Embden J. D., Emmrich F. Enumeration of T cells reactive with Mycobacterium tuberculosis organisms and specific for the recombinant mycobacterial 64-kDa protein. Eur J Immunol. 1987 Mar;17(3):351–357. doi: 10.1002/eji.1830170308. [DOI] [PubMed] [Google Scholar]
  6. Kieser T., Moss M. T., Dale J. W., Hopwood D. A. Cloning and expression of Mycobacterium bovis BCG DNA in "Streptomyces lividans". J Bacteriol. 1986 Oct;168(1):72–80. doi: 10.1128/jb.168.1.72-80.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Kingston A. E., Salgame P. R., Mitchison N. A., Colston M. J. Immunological activity of a 14-kilodalton recombinant protein of Mycobacterium tuberculosis H37Rv. Infect Immun. 1987 Dec;55(12):3149–3154. doi: 10.1128/iai.55.12.3149-3154.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  9. Lamb F. I., Colston M. J. Cloning of Mycobacterium leprae genes in Streptomyces. Lepr Rev. 1986 Dec;57 (Suppl 2):225–229. [PubMed] [Google Scholar]
  10. Lu M. C., Lien M. H., Becker R. E., Heine H. C., Buggs A. M., Lipovsek D., Gupta R., Robbins P. W., Grosskinsky C. M., Hubbard S. C. Genes for immunodominant protein antigens are highly homologous in Mycobacterium tuberculosis, Mycobacterium africanum, and the vaccine strain Mycobacterium bovis BCG. Infect Immun. 1987 Oct;55(10):2378–2382. doi: 10.1128/iai.55.10.2378-2382.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Mehra V., Sweetser D., Young R. A. Efficient mapping of protein antigenic determinants. Proc Natl Acad Sci U S A. 1986 Sep;83(18):7013–7017. doi: 10.1073/pnas.83.18.7013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Mustafa A. S., Gill H. K., Nerland A., Britton W. J., Mehra V., Bloom B. R., Young R. A., Godal T. Human T-cell clones recognize a major M. leprae protein antigen expressed in E. coli. Nature. 1986 Jan 2;319(6048):63–66. doi: 10.1038/319063a0. [DOI] [PubMed] [Google Scholar]
  13. Oftung F., Mustafa A. S., Husson R., Young R. A., Godal T. Human T cell clones recognize two abundant Mycobacterium tuberculosis protein antigens expressed in Escherichia coli. J Immunol. 1987 Feb 1;138(3):927–931. [PubMed] [Google Scholar]
  14. Ottenhoff T. H., Klatser P. R., Ivanyi J., Elferink D. G., de Wit M. Y., de Vries R. R. Mycobacterium leprae-specific protein antigens defined by cloned human helper T cells. Nature. 1986 Jan 2;319(6048):66–68. doi: 10.1038/319066a0. [DOI] [PubMed] [Google Scholar]
  15. Shepard C. C., van Landingham R., Walker L. L. Searches among mycobacterial cultures for antileprosy vaccines. Infect Immun. 1980 Sep;29(3):1034–1039. doi: 10.1128/iai.29.3.1034-1039.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Shinnick T. M., Sweetser D., Thole J., van Embden J., Young R. A. The etiologic agents of leprosy and tuberculosis share an immunoreactive protein antigen with the vaccine strain Mycobacterium bovis BCG. Infect Immun. 1987 Aug;55(8):1932–1935. doi: 10.1128/iai.55.8.1932-1935.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Shinnick T. M. The 65-kilodalton antigen of Mycobacterium tuberculosis. J Bacteriol. 1987 Mar;169(3):1080–1088. doi: 10.1128/jb.169.3.1080-1088.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Thole J. E., Dauwerse H. G., Das P. K., Groothuis D. G., Schouls L. M., van Embden J. D. Cloning of Mycobacterium bovis BCG DNA and expression of antigens in Escherichia coli. Infect Immun. 1985 Dec;50(3):800–806. doi: 10.1128/iai.50.3.800-806.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Thole J. E., Keulen W. J., De Bruyn J., Kolk A. H., Groothuis D. G., Berwald L. G., Tiesjema R. H., van Embden J. D. Characterization, sequence determination, and immunogenicity of a 64-kilodalton protein of Mycobacterium bovis BCG expressed in escherichia coli K-12. Infect Immun. 1987 Jun;55(6):1466–1475. doi: 10.1128/iai.55.6.1466-1475.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  22. Young D. B., Kent L., Young R. A. Screening of a recombinant mycobacterial DNA library with polyclonal antiserum and molecular weight analysis of expressed antigens. Infect Immun. 1987 Jun;55(6):1421–1425. doi: 10.1128/iai.55.6.1421-1425.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Young R. A., Mehra V., Sweetser D., Buchanan T., Clark-Curtiss J., Davis R. W., Bloom B. R. Genes for the major protein antigens of the leprosy parasite Mycobacterium leprae. Nature. 1985 Aug 1;316(6027):450–452. doi: 10.1038/316450a0. [DOI] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES