Decreased Capacity of Recombinant 45/47-kDa Molecules (Apa) ofMycobacterium tuberculosis to Stimulate T Lymphocyte Responses Related to Changes in Their Mannosylation Pattern

• Published in: The Journal of biological chemistry 1999-11, Vol.274 (45), p.32023
• Main Authors: Cynthia Horn, Abdelkader Namane, Pascale Pescher, Michel Rivière, Félix Romain, Germain Puzo, Octavian Bârzu, Gilles Marchal
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.274.45.32023

The Apa molecules secreted by Mycobacterium tuberculosis , Mycobacterium bovis, or BCG have been identified as major immunodominant antigens. Mass spectrometry analysis indicated similar mannosylation, a complete pattern from 1 up to 9 hexose residues/mole of protein, of the native species from the 3 reference strains. The recombinant antigen expressed in M. smegmatis revealed a different mannosylation pattern: species containing 7 to 9 sugar residues/mole of protein were in the highest proportion, whereas species bearing a low number of sugar residues were almost absent. The 45/47-kDa recombinant antigen expressed in E. coli was devoid of sugar residues. The proteins purified from M. tuberculosis, M. bovis, or BCG have a high capacity to elicit in vivo potent delayed-type hypersensitivity (DTH) reactions and to stimulate in vitro sensitized T lymphocytes of guinea pigs immunized with living BCG. The recombinant Apa expressed in Mycobacterium smegmatis was 4-fold less potent in vivo in the DTH assay and 10-fold less active in vitro to stimulate sensitized T lymphocytes than the native proteins. The recombinant protein expressed in Escherichia coli was nearly unable to elicit DTH reactions in vivo or to stimulate T lymphocytes in vitro . Thus the observed biological effects were related to the extent of glycosylation of the antigen.