Isoniazid activation defects in recombinant Mycobacterium tuberculosis catalase-peroxidase (KatG) mutants evident in InhA inhibitor production

Chih Jen Wei, Benfang Lei, James M. Musser, Shiao Chun Tu

Research output: Contribution to journalArticle

68 Scopus citations

Abstract

Mycobacterium tuberculosis KatG catalyzes the activation of the antitubercular agent isoniazid to yield an inhibitor targeting enoyl reductase (InhA). However, no firm biochemical link between many KatG variants and isoniazid resistance has been established. In the present study, six distinct KatG variants identified in clinical Mycobacterium tuberculosis isolates resistant to isoniazid were generated by site-directed mutagenesis, and the recombinant mutant proteins (KatGA110V, KatGA139P, KatGS315N, KatGL619P, KatGL634F, and KatGD735A) were purified and characterized with respect to their catalase-peroxidase activities (in terms of kcat/Km) rates of free-radical formation from isoniazid oxidation, and, moreover, abilities to activate isoniazid. The A110V amino acid replacement did not result in significant alteration of KatG activities except that the peroxidase activity was enhanced. The other mutations, however, resulted in modestly reduced catalase and peroxidase catalytic efficiencies and, for the four mutants tested, significantly lower activities to oxidize isoniazid. Compared to the wild-type enzyme, the ability of the KatGL634F, KatGA139P, and KatGD735A variants to activate isoniazid decreased by 36%, 76%, and 73%, respectively, whereas the KatGS315N and KatGL619P variants completely lost their abilities to convert isoniazid into the InhA inhibitor. In addition, the inclusion of exogenous Mn2+ to the isoniazid activation reaction mix significantly improved the ability of wild-type and KatG mutants to produce the InhA inhibitor.

Original languageEnglish (US)
Pages (from-to)670-675
Number of pages6
JournalAntimicrobial Agents and Chemotherapy
Volume47
Issue number2
DOIs
StatePublished - Feb 1 2003

ASJC Scopus subject areas

  • Pharmacology
  • Pharmacology (medical)
  • Infectious Diseases

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