title: Three-dimensional model and molecular mechanism of Mycobacterium tuberculosis catalase-peroxidase (KatG) and isoniazid-resistant KatG mutants.
abstract: Mycobacterium tuberculosis KatG enzyme functions both as catalase for removing hydrogen peroxide (H(2)O(2)) and as peroxidase for oxidating isoniazid (INH) to active form of anti-tuberculosis drug. Although mutations in M. tuberculosis KatG confer INH resistance in tuberculous patients, structural bases for INH-resistant mutations in the KatG gene remains poorly understood. Here, three M. tuberculosis KatG mutants bearing Arg418--> Gln, Ser315 --> Thr, or Trp321 --> Gly replacement were assessed for changes in catalase-peroxidase activities and possible structure bases relevant to such changes. These three M. tuberculosis KatG mutants exhibited a marked impairment or loss of catalase-peroxidase activities. The possible structural bases for the mutant-induced loss of enzyme activities were then analyzed using a three-dimensional model of M. tuberculosis KatG protein constructed on the basis of the crystal structure of the catalase-peroxidase from Burkholderia pseudomallei. The model suggests that three M. tuberculosis KatG mutants bearing Arg418 --> Gln, Ser315 -->Thr, or Trp321--> Gly replacement affect enzyme activities by different mechanisms, although each of them impacts consequently on a heme-associated structure, the putative oxidative site. Moreover, in addition to the widely accepted substrate-binding site, M. tuberculosis KatG may bear another H(2)O(2) binding site. This H(2)O(2) binding site appears to interact with the catalytic site by a possible electron-transfer chain, a Met255-Tyr229-Trp107 triad conserved in many catalase-peroxidases. The Ser315 --> Thr mutant may have direct effect on the catalytic site by interfering with electron transfer in addition to the previously proposed mechanism of steric constraint.
authors.affiliation: State Key Laboratory of Genetic Engineering, Institute of Genetics, and Huashan Hospital, Fudan University, Shanghai, China 200433.
authors.forename: L
authors.initials: L
authors.lastname: Mo
authors.forename: W
authors.initials: W
authors.lastname: Zhang
authors.forename: J
authors.initials: J
authors.lastname: Wang
authors.forename: X H
authors.initials: XH
authors.lastname: Weng
authors.forename: S
authors.initials: S
authors.lastname: Chen
authors.forename: L Y
authors.initials: LY
authors.lastname: Shao
authors.forename: M Y
authors.initials: MY
authors.lastname: Pang
authors.forename: Z W
authors.initials: ZW
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D002374:Catalase / Q000737:chemistry* / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D007538:Isoniazid / Q000494:pharmacology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D010084:Oxidation-Reduction; D011487:Protein Conformation
chemical_list: D001426:Bacterial Proteins; D002374:Catalase; C480353:katG protein, Mycobacterium tuberculosis; D007538:Isoniazid