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Mycobacterium leprae produces extracellular homologs of the antigen 85 complex
1Department of Microbiology, Colorado State University, Fort Collins 80523.
Abstract:
The antigen 85 complex is a set of at least three closely related secreted proteins (85A, 85B, and 85C) of 30 to 32 kDa produced by Mycobacterium tuberculosis and other mycobacteria. Their prominence in Mycobacterium leprae, the one obligate intracellular pathogen of the genus, had been assumed on the basis of immunological evidence and proof of the existence of the gene encoding the 85B protein of the complex. We have now observed the production of this family of proteins by M. leprae through analysis of various fractions by Western blotting (immunoblotting) with monospecific rabbit antisera raised against the individual Mycobacterium bovis BCG 85A, 85B, and 85C proteins. A predominant cross-reactive band with an apparent molecular mass of 30 kDa was detected in extracts of nondisrupted whole M. leprae and in soluble fractions prepared from the tissues of M. leprae-infected armadillos. Further studies of the subcellular distribution of this protein within the bacterium confirmed that it is secreted by the organism, an observation that explains past difficulties in detecting the antigen 85 complex in M. leprae. Confirmation that the M. leprae product is a member of the antigen 85 complex was obtained by comparison of peptide fingerprints with those from the BCG product. The pattern of reactivity of the M. leprae antigen 85 complex with anti-M. bovis BCG 85B serum, as well as two-dimensional electrophoresis, established that the 85B component was the predominant member of the complex in M. leprae. The fibronectin-binding capacity of the M. leprae and BCG 85 complexes was reinvestigated by new approaches and is questioned. Nevertheless, the results obtained with the native proteins reinforce previous reports, derived primarily from the use of homologous proteins, that the antigen 85 complex is one of the dominant protein immunogens of the leprosy bacillus.
Insights
Researchers confirmed the antigen 85 complex proteins are produced by Mycobacterium leprae. The 85B component is the most abundant, explaining its role as a key immunogen in leprosy.
Area of Science:
- Microbiology
- Immunology
- Molecular Biology
Background:
- The antigen 85 complex (85A, 85B, 85C) are secreted proteins from Mycobacterium species.
- Previous studies suggested the presence of antigen 85 complex in Mycobacterium leprae based on immunological data and gene evidence.
- The obligate intracellular nature of Mycobacterium leprae historically complicated direct detection of its secreted proteins.
Purpose of the Study:
- To directly observe and confirm the production of antigen 85 complex proteins by Mycobacterium leprae.
- To identify the predominant component of the antigen 85 complex within Mycobacterium leprae.
- To investigate the secretion and subcellular localization of these proteins in Mycobacterium leprae.
Main Methods:
- Western blotting using monospecific antisera against Mycobacterium bovis BCG 85A, 85B, and 85C proteins.
- Analysis of various M. leprae fractions, including extracts from infected armadillo tissues.
- Peptide fingerprinting and two-dimensional electrophoresis for protein identification and comparison.
- Subcellular fractionation to determine protein localization.
Main Results:
- A predominant 30 kDa cross-reactive protein band, indicative of the antigen 85 complex, was detected in M. leprae extracts and soluble fractions.
- The protein was confirmed to be secreted by M. leprae, resolving previous detection challenges.
- Peptide fingerprinting confirmed the M. leprae product belongs to the antigen 85 complex.
- Two-dimensional electrophoresis and reactivity with anti-M. bovis BCG 85B serum identified the 85B component as the predominant member.
Conclusions:
- Mycobacterium leprae actively produces the antigen 85 complex, with the 85B component being the most abundant.
- The secretion of this complex explains prior difficulties in its detection in M. leprae.
- The antigen 85 complex, particularly the 85B component, is a dominant protein immunogen of the leprosy bacillus.