CATALASE-PEROXIDASE CATALYSIS IN ANTIBIOTIC ACTIVATION

描述：(改编自申请者的摘要) 提出研究是为了解释分枝杆菌的催化功能 结核过氧化氢酶，一种血红素酶，在激活 抗分枝杆菌抗生素异烟肼(异烟酸 酰肼)。具体目标包括识别近端 过氧化氢酶-过氧化物酶中血红素铁的配基，自旋的表征 休眠酶中血红素铁的状态和配位数， 高价酶中间体的鉴定及动力学分析 这些中间体与异烟肼的反应。特别之处 令人感兴趣的是氧亚铁过氧化氢酶的潜在催化能力- 过氧化物酶，以及过氧亚硝酸盐激活该酶的潜力。 选择的氨基酸残基在催化机理和 在异烟肼结合中将通过检查 两种突变型过氧化氢酶-过氧化物酶的性质 临床分离的耐异烟肼结核分枝杆菌菌株。 另一种结核分枝杆菌酶--脂肪酰烯醇酰的抑制作用 还原酶(Inha蛋白)被认为是药物作用的靶点，通过 异烟肼激活的过氧化氢酶-过氧化物酶，也将被研究。 其他目的是解决过氧化氢酶过氧化物酶的锰(II)过氧化物酶活性 被认为是重要的，因为Mn(III)是一个有效的单电子 异烟肼的氧化剂。光截流技术 光谱学、共振拉曼和电子顺磁共振 光谱学将应用于实验方案。这个 拟议研究的结果将促进对 当前使用的一线抗生素在治疗中的作用 肺结核。

DESCRIPTION: (adapted from applicant's abstract) The major role of the propose research is to explain the catalytic function of Mycobacterium Tuberculosis catalase-peroxidase, a heme-enzyme, in the activation of the antimycobacterial antibiotic, isoniazid (isonicotinic acid hydrazide). The specific aims include identification of the proximal ligand to heme iron in catalase-peroxidase, characterization of the spin state and coordination number of the heme iron in the resting enzyme, identification of hypervalent enzyme intermediates, and kinetic analysis of the reaction of these intermediates with isoniazid. Of special interest is the potential catalytic competence of oxy-ferrous catalase- peroxidase, and the potential for peroxynitrite to activate the enzyme. The role of selected amino acid residues in the catalytic mechanism an in isoniazid binding will be evaluated through examination of the properties of two mutant catalase-peroxidase enzymes identified from clinically isolated, isoniazid resistant M. tuberculosis strains. Inhibition of another M. tuberculosis enzyme, a fatty acyl enoyl reductase (the inhA protein) thought to be a target of drug action, by isoniazid activated catalase-peroxidase, will also be investigated. Other aims address the Mn(II)-peroxidase activity of catalase-peroxidase considered important because Mn(III) is an efficient single electron oxidant of isoniazid. The techniques of optical stopped-flow spectroscopy, resonance Raman, and electron paramagnetic resonance spectroscopies will be applied in the experimental protocols. The results of the proposed studies will advance a detailed understanding of the action of a first line antibiotic in current use to treat tuberculosis.