Hydrogen Peroxide-Mediated Isoniazid Activation Catalyzed by Mycobacterium tuberculosis Catalase-Peroxidase (KatG) and Its S315T Mutant

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• 作者：Xiangbo Zhao ; Hong Yu ; Shengwei Yu ; Feng Wang ; James C. Sacchettini ; and Richard S. Magliozzo
• 刊名：Biochemistry
• 出版年：2006
• 出版时间：April 4, 2006
• 年：2006
• 卷：45
• 期：13
• 页码：4131 - 4140
• 全文大小：173K
• 年卷期：v.45,no.13(April 4, 2006)
• ISSN：1520-4995

文摘

Inhibition of the enzyme Mycobacterium tuberculosis InhA (enoyl-acyl carrier protein reductase)due to formation of an isonicotinoyl-NAD adduct (IN-NAD) from isoniazid (INH) and nicotinamide adeninedinucleotide cofactor is considered central to the mode of action of INH, a first-line treatment fortuberculosis infection. INH action against mycobacteria requires catalase-peroxidase (KatG) function,and IN-NAD adduct formation is catalyzed in vitro by M. tuberculosis KatG under a variety of conditions,yet a physiologically relevant approach to the process has not emerged that allows scrutiny of the mechanismand the origins of INH resistance in the most prevalent drug-resistant strain bearing KatG[S315T]. In thisreport, we describe how hydrogen peroxide, delivered at very low concentrations to ferric KatG, leads toefficient inhibition of InhA due to formation of the IN-NAD adduct. The rate of adduct formation mediatedby wild-type KatG was about 20-fold greater than by the isoniazid-resistant KatG[S315T] mutant underoptimal conditions (H₂O₂ supplied along with NAD⁺ and INH). Slow adduct formation also occurs startingwith NADH and INH, in the presence of KatG even in the absence of added peroxide, due to endogenousperoxide. The poor efficiency of the KatG[S315T] mutant can be enhanced merely by increasing theconcentration of INH, consistent with this enzyme's reduced affinity for INH binding to the resting enzymeand the catalytically competent enzyme intermediate (Compound I). Origins of drug resistance in theKatG[S315T] mutant enzyme are analyzed at the structural level through examination of the three-dimensional X-ray crystal structure of the mutant enzyme.