SITE-DIRECTED MUTAGENESIS OF THE KATG GENE OF MYCOBACTERIUM-TUBERCULOSIS - EFFECTS ON CATALASE-PEROXIDASE ACTIVITIES AND ISONIAZID RESISTANCE

Recent studies examining the molecular mechanisms of isoniazid (INH) r esistance in Mycobacterium tuberculosis have demonstrated that a signi ficant percentage of drug-resistant strains are mutated in the katG ge ne which encodes a catalase-peroxidase, and the majority of these alte rations are missense mutations which result in the substitution of a s ingle amino acid. In previous reports, residues which may be critical for enzymatic activity and the drug-resistant phenotype have been iden tified by evaluating INH-resistant clinical isolates and in vitro muta nts. In this study, site-directed mutagenesis techniques were utilized to alter the wild-type katG gene from M. tuberculosis at 13 of these codons. The effects of these mutations were determined using complemen tation assays in katG-defective, INH-resistant strains of Mycobacteriu m smegmatis and Mycobacterium bovis BCG. This mutational analysis reve aled that point mutations in the katG gene at nine of the 13 codons ca n cause drug resistance, and that enzymatic activity and resistance to INH are inversely related. In addition, mutations in the mycobacteria l catalase-peroxidase which reduce catalase activity also decrease per oxidase activity.