The isoniazid analog IQG-607 is not a direct substrate for the Mycobacterium tuberculosis catalase-peroxidase KatG

Abstract

Tuberculosis (TB) is an infectious disease caused mainly by Mycobacterium tuberculosis (Mtb) and is responsible for millions of deaths. New Mtb strains resistant to TB drugs are emerging and spreading. The first-line TB drug, isoniazid (INH), must be activated inside mycobacterial cells by the catalase-peroxidase enzyme KatG to exert its antimicrobial activity, and mutations on the katG gene are a significant cause of INH resistance in clinics. The metal-containing compound IQG-607 is an INH analog developed to inhibit the target of INH, the FASII enzyme enoyl-ACP-reductase (InhA), without requiring KatG. However, we recently showed that inside mycobacterial cells, IQG-607 activity depends on KatG. Hence, this compound might also be activated by KatG to inhibit InhA. We evaluated whether recombinant MtKatG uses IQG-607 as a substrate in oxidation reactions and adduct formation with NAD+. A recombinant MtKatG was produced in E. coli and purified in a 3-step protocol to obtain a homogeneous protein. An HPLC method was optimized to monitor both oxidation and adduct products, and our assay system was validated by performing control reactions using INH as a substrate. We found that the metal-based compound IQG-607 is not a substrate for recombinant MtKatG under all conditions tested.