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. 1997 Jan;65(1):267–271. doi: 10.1128/iai.65.1.267-271.1997

Delayed-type hypersensitivity activity of the Brucella L7/L12 ribosomal protein depends on posttranslational modification.

G Bachrach 1, M Banai 1, Y Fishman 1, H Bercovier 1
PMCID: PMC174586  PMID: 8975922

Abstract

The ribosomal protein L7/L12 isolated from Brucella melitensis induces a delayed-type hypersensitivity (DTH) reaction in brucella-sensitized guinea pigs. Surprisingly, the recombinant brucella L7/L12 protein expressed in Escherichia coli as a fusion protein with a six-histidine tag cannot elicit such a reaction. The six histidines tagged to the recombinant L7/L12 protein were removed enzymatically, but the resulting protein did not induce a DTH reaction in sensitized animals. Incubation of the recombinant L7/L12 fusion protein in a B. melitensis lysate endowed the recombinant protein with a DTH activity, suggesting that the recombinant protein was modified by this treatment. Glycosylation or phosphorylation of the recombinant L7/L12 protein could not be detected. On the other hand, radiolabeled palmitic acid was found to be incorporated to the recombinant protein during its incubation in the brucella lysate. This incorporation was specific for the brucella L7/L12 protein and was inhibited when the brucella lysate was frozen and thawed before the incubation. The data reported here indicate that posttranslational modification of L7/L12 protein comprising at least an acylation step is required for the brucella L7/L12 DTH activity.

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Selected References

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  1. Abshire K. Z., Neidhardt F. C. Growth rate paradox of Salmonella typhimurium within host macrophages. J Bacteriol. 1993 Jun;175(12):3744–3748. doi: 10.1128/jb.175.12.3744-3748.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ashbridge K. R., Bäckström B. T., Liu H. X., Vikerfors T., Englebretsen D. R., Harding D. R., Watson J. D. Mapping of T helper cell epitopes by using peptides spanning the 19-kDa protein of Mycobacterium tuberculosis. Evidence for unique and shared epitopes in the stimulation of antibody and delayed-type hypersensitivity responses. J Immunol. 1992 Apr 1;148(7):2248–2255. [PubMed] [Google Scholar]
  3. Bachrach G., Banai M., Bardenstein S., Hoida G., Genizi A., Bercovier H. Brucella ribosomal protein L7/L12 is a major component in the antigenicity of brucellin INRA for delayed-type hypersensitivity in brucella-sensitized guinea pigs. Infect Immun. 1994 Dec;62(12):5361–5366. doi: 10.1128/iai.62.12.5361-5366.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bachrach G., Bar-Nir D., Banai M., Bercovier H. Identification and nucleotide sequence of Brucella melitensis L7/L12 ribosomal protein. FEMS Microbiol Lett. 1994 Jul 15;120(3):237–240. doi: 10.1111/j.1574-6968.1994.tb07039.x. [DOI] [PubMed] [Google Scholar]
  5. Baker R. E., Hill W. E., Larson C. L. Ribosomes of acid-fast bacilli: immunogenicity, serology, and in vitro correlates of delayed hypersensitivity. Infect Immun. 1973 Aug;8(2):236–244. doi: 10.1128/iai.8.2.236-244.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bessler W. G., Cox M., Lex A., Suhr B., Wiesmüller K. H., Jung G. Synthetic lipopeptide analogs of bacterial lipoprotein are potent polyclonal activators for murine B lymphocytes. J Immunol. 1985 Sep;135(3):1900–1905. [PubMed] [Google Scholar]
  7. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  8. Brooks-Worrell B. M., Splitter G. A. Antigens of Brucella abortus S19 immunodominant for bovine lymphocytes as identified by one- and two-dimensional cellular immunoblotting. Infect Immun. 1992 Jun;60(6):2459–2464. doi: 10.1128/iai.60.6.2459-2464.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Corbel M. J. The immunogenic activity of ribosomal fractions derived from Brucella abortus. J Hyg (Lond) 1976 Feb;76(1):65–74. doi: 10.1017/s0022172400054954. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dannenberg A. M., Jr Roles of cytotoxic delayed-type hypersensitivity and macrophage-activating cell-mediated immunity in the pathogenesis of tuberculosis. Immunobiology. 1994 Oct;191(4-5):461–473. doi: 10.1016/S0171-2985(11)80452-3. [DOI] [PubMed] [Google Scholar]
  11. Dobos K. M., Swiderek K., Khoo K. H., Brennan P. J., Belisle J. T. Evidence for glycosylation sites on the 45-kilodalton glycoprotein of Mycobacterium tuberculosis. Infect Immun. 1995 Aug;63(8):2846–2853. doi: 10.1128/iai.63.8.2846-2853.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Espitia C., Mancilla R. Identification, isolation and partial characterization of Mycobacterium tuberculosis glycoprotein antigens. Clin Exp Immunol. 1989 Sep;77(3):378–383. [PMC free article] [PubMed] [Google Scholar]
  13. Garbe T., Harris D., Vordermeier M., Lathigra R., Ivanyi J., Young D. Expression of the Mycobacterium tuberculosis 19-kilodalton antigen in Mycobacterium smegmatis: immunological analysis and evidence of glycosylation. Infect Immun. 1993 Jan;61(1):260–267. doi: 10.1128/iai.61.1.260-267.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gerardy-Schahn R., Bethe A., Brennecke T., Mühlenhoff M., Eckhardt M., Ziesing S., Lottspeich F., Frosch M. Molecular cloning and functional expression of bacteriophage PK1E-encoded endoneuraminidase Endo NE. Mol Microbiol. 1995 May;16(3):441–450. doi: 10.1111/j.1365-2958.1995.tb02409.x. [DOI] [PubMed] [Google Scholar]
  15. Gregory R. L. Microbial ribosomal vaccines. Rev Infect Dis. 1986 Mar-Apr;8(2):208–217. doi: 10.1093/clinids/8.2.208. [DOI] [PubMed] [Google Scholar]
  16. Hamburger J., Zuckerman A. Plasmodium berghei: I. Immunochemical properties of fractions of a soluble extract. Exp Parasitol. 1976 Jun;39(3):460–478. doi: 10.1016/0014-4894(76)90050-3. [DOI] [PubMed] [Google Scholar]
  17. Hauschildt S., Hoffmann P., Beuscher H. U., Dufhues G., Heinrich P., Wiesmüller K. H., Jung G., Bessler W. G. Activation of bone marrow-derived mouse macrophages by bacterial lipopeptide: cytokine production, phagocytosis and Ia expression. Eur J Immunol. 1990 Jan;20(1):63–68. doi: 10.1002/eji.1830200110. [DOI] [PubMed] [Google Scholar]
  18. Issartel J. P., Koronakis V., Hughes C. Activation of Escherichia coli prohaemolysin to the mature toxin by acyl carrier protein-dependent fatty acylation. Nature. 1991 Jun 27;351(6329):759–761. doi: 10.1038/351759a0. [DOI] [PubMed] [Google Scholar]
  19. Jones L. M., Berman D. T. Antibody-mediated and delayed-type hypersensitivity reactions to Brucella skin test antigens in guinea pigs. Infect Immun. 1975 Feb;11(2):360–364. doi: 10.1128/iai.11.2.360-364.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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]
  21. Loge R. V., Hill W. E., Baker R. E., Larson C. L. Delayed hypersensitivity reactions provoked by ribosomes from acid-fast bacilli: physical characteristics and immunological aspects of core ribosomal proteins from Mycobacterium smegmatis. Infect Immun. 1974 Mar;9(3):489–496. doi: 10.1128/iai.9.3.489-496.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Oliveira S. C., Zhu Y., Splitter G. A. Recombinant L7/L12 ribosomal protein and gamma-irradiated Brucella abortus induce a T-helper 1 subset response from murine CD4+ T cells. Immunology. 1994 Dec;83(4):659–664. [PMC free article] [PubMed] [Google Scholar]
  23. Orme I. M., Miller E. S., Roberts A. D., Furney S. K., Griffin J. P., Dobos K. M., Chi D., Rivoire B., Brennan P. J. T lymphocytes mediating protection and cellular cytolysis during the course of Mycobacterium tuberculosis infection. Evidence for different kinetics and recognition of a wide spectrum of protein antigens. J Immunol. 1992 Jan 1;148(1):189–196. [PubMed] [Google Scholar]
  24. Ortiz-Ortiz L., Solarolo E. B., Bojalil L. F. Delayed hypersensitivity to ribosomal protein from BCG. J Immunol. 1971 Oct;107(4):1022–1026. [PubMed] [Google Scholar]
  25. Schmidt M. F. Fatty acylation of proteins. Biochim Biophys Acta. 1989 Dec 6;988(3):411–426. doi: 10.1016/0304-4157(89)90013-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Tantimavanich S., Nagai S., Nomaguchi H., Kinomoto M., Ohara N., Yamada T. Immunological properties of ribosomal proteins from Mycobacterium bovis BCG. Infect Immun. 1993 Sep;61(9):4005–4007. doi: 10.1128/iai.61.9.4005-4007.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Terhorst C., Wittmann-Liebold B., Möller W. 50-S ribosomal proteins. Peptide studies on two acidic proteins, A 1 and A 2 , isolated from 50-S ribosomes of Escherichia coli. Eur J Biochem. 1972 Jan 31;25(1):13–19. doi: 10.1111/j.1432-1033.1972.tb01661.x. [DOI] [PubMed] [Google Scholar]
  28. Thompson H. C., Snyder I. S. Protection against pneumococcal infection by a ribosomal preparation. Infect Immun. 1971 Jan;3(1):16–23. doi: 10.1128/iai.3.1.16-23.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Venneman M. R., Bigley N. J. Isolation and partial characterization of an immunogenic moiety obtained from Salmonella typhimurium. J Bacteriol. 1969 Oct;100(1):140–148. doi: 10.1128/jb.100.1.140-148.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wang X. H., Ohmen J. D., Uyemura K., Rea T. H., Kronenberg M., Modlin R. L. Selection of T lymphocytes bearing limited T-cell receptor beta chains in the response to a human pathogen. Proc Natl Acad Sci U S A. 1993 Jan 1;90(1):188–192. doi: 10.1073/pnas.90.1.188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. YOUMANS A. S., YOUMANS G. P. IMMUNOGENIC ACTIVITY OF A RIBOSOMAL FRACTION OBTAINED FROM MYCOBACTERIUM TUBERCULOSIS. J Bacteriol. 1965 May;89:1291–1298. doi: 10.1128/jb.89.5.1291-1298.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]

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