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. 1993 Jun 1;90(11):5322–5326. doi: 10.1073/pnas.90.11.5322

Isolation of a proline-rich mycobacterial protein eliciting delayed-type hypersensitivity reactions only in guinea pigs immunized with living mycobacteria.

F Romain 1, J Augier 1, P Pescher 1, G Marchal 1
PMCID: PMC46708  PMID: 8506381

Abstract

Effective protection against a virulent challenge with Mycobacterium tuberculosis is induced only by a previous immunization with living attenuated mycobacteria, usually bacillus Calmette-Guérin (BCG). Living and killed bacteria share a number of common antigens. To identify and to purify molecules that are dominant antigens during immunization with living bacteria, a two-step selection procedure was used. Quantitative delayed-type hypersensitivity (DTH) reactions elicited in guinea pigs immunized either with living or with killed BCG were used to select or counterselect antigens present in BCG culture filtrates. Each major fraction eluted from a series of HPLC columns (gel filtration, DEAE, reverse-phase chromatography) was assayed and titrated on guinea pigs of each group. A protein with an unusual amino acid composition (40% proline, 12% threonine) was purified and N-terminally sequenced. To our knowledge, the sequence Thr-Pro-Pro-Xaa-Glu-Xaa-Pro-Pro-Pro-Pro-Gln-Xaa-Val-Xaa-Leu has not been previously reported. The protein was 100-fold more potent on guinea pigs immunized with living bacteria than on guinea pigs immunized with dead bacteria to elicit a DTH reaction.

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Selected References

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

  1. Augier J., Augier-Gibory S., Romain F., Versmisse E. Search of a standard for the titration of antigen L. Dev Biol Stand. 1986;58(Pt B):495–501. [PubMed] [Google Scholar]
  2. Caulada-Benedetti Z., al-Zamel F., Sher A., James S. Comparison of Th1- and Th2-associated immune reactivities stimulated by single versus multiple vaccination of mice with irradiated Schistosoma mansoni cercariae. J Immunol. 1991 Mar 1;146(5):1655–1660. [PubMed] [Google Scholar]
  3. Coon J., Hunter R. Selective induction of delayed hypersensitivity by a lipid conjugated protein antigen which is localized in thymus dependent lymphoid tissue. J Immunol. 1973 Jan;110(1):183–190. [PubMed] [Google Scholar]
  4. DUBOS R. J., PIERCE C. H., SCHAEFER W. B. Antituberculous immunity induced in mice by vaccination with living cultures of attenuated tubercle bacilli. J Exp Med. 1953 Feb 1;97(2):207–220. doi: 10.1084/jem.97.2.207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Daynes R. A., Araneo B. A., Dowell T. A., Huang K., Dudley D. Regulation of murine lymphokine production in vivo. III. The lymphoid tissue microenvironment exerts regulatory influences over T helper cell function. J Exp Med. 1990 Apr 1;171(4):979–996. doi: 10.1084/jem.171.4.979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Furthmayr H., Timpl R. Characterization of collagen peptides by sodium dodecylsulfate-polyacrylamide electrophoresis. Anal Biochem. 1971 Jun;41(2):510–516. doi: 10.1016/0003-2697(71)90173-4. [DOI] [PubMed] [Google Scholar]
  7. Gajewski T. F., Schell S. R., Nau G., Fitch F. W. Regulation of T-cell activation: differences among T-cell subsets. Immunol Rev. 1989 Oct;111:79–110. doi: 10.1111/j.1600-065x.1989.tb00543.x. [DOI] [PubMed] [Google Scholar]
  8. Kanai K. Experimental studies on host-parasite equilibrium in tuberculous infection, in relation to vaccination and chemotherapy. Jpn J Med Sci Biol. 1966 Aug;19(4):181–199. doi: 10.7883/yoken1952.19.181. [DOI] [PubMed] [Google Scholar]
  9. 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]
  10. Landi S., Held H. R., Gupta K. C. The multi-facets of tuberculin standardization. Dev Biol Stand. 1975;29:393–411. [PubMed] [Google Scholar]
  11. Marchal G., Seman M., Milon G., Truffa-Bachi P., Zilberfarb V. Local adoptive transfer of skin delayed-type hypersensitivity initiated by a single T lymphocyte. J Immunol. 1982 Sep;129(3):954–958. [PubMed] [Google Scholar]
  12. Orme I. M. Induction of nonspecific acquired resistance and delayed-type hypersensitivity, but not specific acquired resistance in mice inoculated with killed mycobacterial vaccines. Infect Immun. 1988 Dec;56(12):3310–3312. doi: 10.1128/iai.56.12.3310-3312.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Pearson W. R., Lipman D. J. Improved tools for biological sequence comparison. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2444–2448. doi: 10.1073/pnas.85.8.2444. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Romain F., Laqueyrerie A., Militzer P., Pescher P., Chavarot P., Lagranderie M., Auregan G., Gheorghiu M., Marchal G. Identification of a Mycobacterium bovis BCG 45/47-kilodalton antigen complex, an immunodominant target for antibody response after immunization with living bacteria. Infect Immun. 1993 Feb;61(2):742–750. doi: 10.1128/iai.61.2.742-750.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Rothbard J. B., Gefter M. L. Interactions between immunogenic peptides and MHC proteins. Annu Rev Immunol. 1991;9:527–565. doi: 10.1146/annurev.iy.09.040191.002523. [DOI] [PubMed] [Google Scholar]
  16. Scott P., Natovitz P., Coffman R. L., Pearce E., Sher A. Immunoregulation of cutaneous leishmaniasis. T cell lines that transfer protective immunity or exacerbation belong to different T helper subsets and respond to distinct parasite antigens. J Exp Med. 1988 Nov 1;168(5):1675–1684. doi: 10.1084/jem.168.5.1675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Townsend A., Ohlén C., Bastin J., Ljunggren H. G., Foster L., Kärre K. Association of class I major histocompatibility heavy and light chains induced by viral peptides. Nature. 1989 Aug 10;340(6233):443–448. doi: 10.1038/340443a0. [DOI] [PubMed] [Google Scholar]
  18. WEISS D. W., DUBOS R. J. Antituberculous immunity induced in mice by vaccination with killed tubercle bacilli or with a soluble bacillary extract. J Exp Med. 1955 Mar 1;101(3):313–330. doi: 10.1084/jem.101.3.313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Young D. B., Kaufmann S. H., Hermans P. W., Thole J. E. Mycobacterial protein antigens: a compilation. Mol Microbiol. 1992 Jan;6(2):133–145. doi: 10.1111/j.1365-2958.1992.tb01994.x. [DOI] [PubMed] [Google Scholar]

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