Structural determinants of the oxidative stress response

这项拟议研究的主要目标是表征保护生物体免受氧化应激的蛋白质。20多年来，Loewen实验室的研究一直集中在单功能过氧化氢酶和双功能过氧化氢酶-过氧化氢酶，它们的主要生理作用是将潜在有毒的过氧化氢降解为无害的水和氧气。从细菌遗传分析，到分子遗传特征到蛋白质结构确定，工作已经取得了进展，并极大地提高了对酶如何响应不同生物体中的生理挑战的理解。最近在实验室采用了蛋白质X射线结晶学程序，这使得解决酶中作用的催化机制的问题成为可能；拟议的研究将继续朝着这个方向进行。拟议研究的目的是确定保护生物体免受过氧化氢伤害的因素，具体目标是确定影响过氧化氢酶-过氧化物酶和单功能过氧化氢酶催化机制的结构特征并使其合理化。作为广泛使用的抗结核药物异烟肼的激活剂，双功能过氧化氢酶-过氧化物酶已经获得了相当大的恶名。特别是，缺乏这种酶的结核分枝杆菌菌株对这种药物具有抗药性，这是耐异烟肼结核病传播的最常见原因，并产生了严重的健康风险。拟议研究的一个分支将侧重于确定异烟肼激活所涉及的催化机制。这项研究将通过更好地了解过氧化氢降解的机制和异烟肼作为抗结核药物的激活，从而增强我们对酶催化的了解。

The primary goal of the proposed research is to characterize proteins that protect organisms against oxidative stress. For over 20 years, research in the Loewen laboratory has been focused on monofunctional catalases and bifunctional catalase-peroxidases which have as their main physiological role the degradation of potentially toxic hydrogen peroxide to harmless water and oxygen. Work has progressed from bacterial genetic analysis, through molecular genetic characterization to protein structure determination and has provided a greatly improved understanding of how the enzymes respond to physiological challenge in different organisms. The recent adoption of protein X-ray crystallography procedures in the laboratory has made it possible to address questions about the catalytic mechanisms operating in the enzymes; the proposed research will continue in this direction. The goal of the proposed research is to characterize factors that protect organisms against hydrogen peroxide and the specific objectives are to characterize and rationalize structural features that affect the catalytic mechanisms in catalase-peroxidases and monofunctional catalases. The bifunctional catalase-peroxidases have gained considerable notoriety from their role as the activator of the widely used anti-tubercular pro-drug isoniazid. In particular, strains of M. tuberculosis lacking the enzyme are resistant to the drug, giving rise to the most common cause of the spread of isoniazid-resistant tuberculosis and producing a serious health risk. One branch of the proposed research will focus on defining the catalytic mechanism involved in isoniazid activation. The research will enhance our knowledge of enzyme catalysis by providing a better understanding of the mechanisms of hydrogen peroxide degradation and of isoniazid activation as a drug against tuberculosis.