The increase in tuberculosis in the United States is associated with both AIDS patients and isoniazid resistance. A single tuberculosis gene katG appears to play a key role in the mechanism of isoniazid resistance. Isonicotinic acid hydrazide (isoniazid, INH) forms the core of antituberculosis regimens. A single tuberculosis gene katG, encoding both catalase and peroxidase activity, appears to be key in the mechanism of action of isoniazid. The peroxidase catalyzed oxidation of isoniazid and the subsequent inactivation of this protein by the resulting free radicals is proposed to be the mode of action of isoniazid. This gene confers isoniazid susceptibility in resistant microorganisms and its absence is associated with isoniazid resistance in patient isolates of M. tuberculosis. Although the metabolic activation of hydrazines to free radicals by peroxidases has received considerable attention, isoniazid has only been studied once. The oxidation of hydrazines by hemoproteins such as peroxidases typically produces free radicals that covalently bind to the prosthetic heme group. These free radical reactions modify or terminate catalytic function. In the case of the katG protein, which is known to protect against oxidative stress, the sudden loss of its catalase and peroxidase activities could cause the death of M. tuberculosis and be the basis of the mode of action of isoniazid. In addition, isoniazid-derived radicals can reduce oxygen ultimately forming superoxide and the hydroxyl radical, and thereby increase oxidative stress. We plan to measure and determine the structure of the free radical metabolites of isoniazid formed by the protein encoded for by the katG gene of M. tuberculosis. The peroxidase and catalase activities of this protein will be measured after treatment with isoniazid for it is anticipated that isoniazid is a suicide substrate.
