Characterization of fibronectin-binding antigens released by Mycobacterium tuberculosis and Mycobacterium bovis BCG

Abstract

Fibronectin (FN)-binding antigens are prominent components of short-term culture supernatants of Mycobacterium tuberculosis. In 3-day-old supernatants, a 30-kilodalton (kDa) protein was identified as the major FN-binding molecule. In 21-day-old supernatants, FN bound to a double protein band of 30 and 31 kDa, as well as to a group of antigens of larger molecular mass (57 to 60 kDa). FN-binding molecules in this size range, but not of 30 to 31 kDa, were also found in sonicates. We showed that the 31- and 30-kDa FN-binding bands correspond to components A and B of the BCG85 complex, previously shown to be abundant in culture supernatants of Mycobacterium bovis BCG. Thus, a polyclonal antibody to the BCG85 complex bound to the 30- and 31-kDa antigens and inhibited binding of FN to them on immunoblots of the culture filtrates. Similarly, FN bound to the purified components of the BCG85 complex, and this binding was blocked by the antibody. A monoclonal antibody, HYT27, also bound both to the BCG85 components A and B and to the 30- and 31-kDa FN-binding molecules of M. tuberculosis, but it did not block the binding of FN. Related molecules appear to be present on the surface of BCG and to mediate the binding of BCG to FN-coated plastic surfaces, since this binding could also be blocked by the polyclonal anti-BCG85 antibody and by the purified components of BCG85, particularly component A, but not by monoclonal antibody HYT27. The binding of these mycobacterial antigens to FN appears to be of very high affinity, and we suggest that this property of major secreted antigens of M. tuberculosis indicates an important role in mycobacterial disease and in the binding of BCG to tumor cells during immunotherapy of bladder cancer.

Immunodiagnosis of tuberculous meningitis: rapid detection of mycobacterial antigens in cerebrospinal fluid by reverse passive hemagglutination assay and their characterization by Western blotting

“…This may be due to physicochemical di¡erences between secreted and intracellular proteins with regard to charge properties [23]. Besides diagnostic importance, 30^32 kDa antigens seem to have an important role in pathogenesis of tuberculosis [24] as that has been known to bind ¢bronectin [25]. 30^32 kDa and 71 kDa antigens have been shown to induce protective immunity in the host [26].…”

“…The sensitivity of RPHA was determined by titrating various concentrations of MTSE (0.062^128 Wg ml 31 ) in 25 Wl volumes in U-bottom microtiter plates (Dynatech, USA) containing 1% BSA in 25 Wl of PBS (diluent). A total of 25 Wl of coated cells were dispensed per well and shaken vigorously for 2 min and hemagglutination patterns scored according to Stavitsky [16] after 2 h of incubation at room temperature.…”

Comparative cell wall core biosynthesis in the mycolated pathogens,Mycobacterium tuberculosisandCorynebacterium diphtheriae

“…Sequence analysis revealed that three catalytic triad residues had all been replaced and, consistent with these substitutions, the purified recombinant protein did not possess a mycolyltransferase activity [191]. This family of antigens also interact with human fibronectin thereby enhancing the uptake of mycobacteria by human macrophages via complement-mediated phagocytosis [192]. However FbpC1 also lacks the proposed fibronectin-binding region [193] and its biological function remains unclear.…”

The ability to form biofilm influences Mycobacterium avium invasion and translocation of bronchial epithelial cells

“…Recent work has demonstrated that M. avium can bind to human bronchial epithelium by interacting with the β 1 integrin, the fibronectin receptor (Middleton et al ., 2000). M. avium , as with many other pathogenic mycobacteria, can bind to fibronectin through the fibronectin attachment protein and the antigen 85 (Abou-Zeid et al ., 1988;Schorey et al ., 1996).…”