The systems participating in detoxification of reactive oxygen intermediates in Mycobacterium tuberculosis are believed to play a dual role in the biology of this highly adapted human pathogen: (i) they may contribute to the survival of this bacterium in the host; and (ii) alterations in the gene encoding catalase/peroxidase have been linked to this organism's resistance to the front-line antituberculosis drug isoniazid. These relationships prompted us to extend investigations of the oxidative-stress-response systems in M. tuberculosis by analysing the alkyl hydroperoxide reductase gene ahpC and its putative regulator oxyR. Surprisingly, the oxyR gene was found to be inactivated by multiple lesions in M. tuberculosis H37Rv. These alterations were observed in all M. tuberculosis strains tested, and in members of the M. tuberculosis complex: Mycobacterium bovis BCG, Mycobacterium africanum, and Mycobacterium microti. The corresponding region carrying these genes in Mycobacterium leprae, an organism not sensitive to isoniazid, has a complete oxyR gene divergently transcribed from ahpC. An increase in minimal inhibitory concentration for isoniazid was observed upon transformation of M. tuberculosis H37Rv with cosmids carrying the oxyR-ahpC region of M. leprae. In keeping with the observed inactivation of oxyR, transcriptional activity of the corresponding region in M. tuberculosis was an order of magnitude lower than that of the oxyR gene from M. leprae. While the loss of this putative regulator of oxidative-stress response in M. tuberculosis is paradoxical considering the fact that survival in host macrophages is regarded as a critical feature of this pathogen, it offers a partial explanation for the exquisite sensitivity of M. tuberculosis to isoniazid.

The gene encoding a 23 kilodalton protein antigen has been cloned from Mycobacterium tuberculosis by screening of a recombinant DNA library with monoclonal antibodies. The product of the gene has been identified as the superoxide dismutase (SOD) of M. tuberculosis on the basis of sequence comparison and by expression of the recombinant protein in a functionally active form. The derived amino acid sequence of M. tuberculosis SOD reveals a close similarity to manganese-containing SODs from other organisms, in spite of the fact that previous studies using the purified enzyme have identified iron as the preferred metal ion ligand. SOD is present in the extracellular fluid of logarithmic-phase cultures of M. tuberculosis, but the structural gene is not preceded by a signal peptide sequence. Insertion of the M. tuberculosis SOD gene into a novel shuttle vector demonstrated the mycobacteria but is ineffective in Escherichia coli.

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Isoniazid-resistant isolates of Mycobacterium tuberculosis were transformed with a plasmid vector carrying the functional catalase-peroxidase (katG) gene. Expression of katG restored full drug susceptibility in isolates initially resistant to concentrations ranging from 3.2 to > 50 micrograms ml-1. Transformation with the corresponding katG gene from Escherichia coli resulted in low-level expression of catalase and peroxidase activities and conferred partial isoniazid sensitivity.