P450-Mediated Dehydrogenation Mechanisms

P450 介导的脱氢机制

基本信息

批准号：
7048271
负责人：
Garold S Yost
金额：
$ 29万
依托单位：
UNIVERSITY OF UTAH
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-01-01 至 2009-12-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Bioactivation of xenobiotics to toxic intermediates through cytochrome P450 oxygenation mechanisms is a well recognized process. However, the production of electrophilic intermediates by several P450 enzymes (e.g. 1A2, 2B6, 2E1, 2F1, 3A4, and 4B1), through dehydrogenation pathways has only recently been investigated, and the mechanisms that govern selective dehydrogenation rather than oxygenation are not established. Several of the dehydrogenated intermediates are so reactive that they inactivate the P450 enzymes, generally through alkylation of active site nucleophilic residues. Research concerning the catalytic behavior of these specific P450 enzymes and their propensity to dehydrogenate rather than oxygenate substrates is vitally needed. The hypothesis of this research is: the unique catalytic mechanism(s) of facilitated electron transport that determines dehydrogenation by certain P450 enzymes results in xenobiotic-mediated injury and altered drug metabolism in humans. The specific goals of this application are to determine the characteristics of the enzyme active-site environment that direct dehydrogenation mechanisms of specific cytochrome P450 enzymes, and to define the substrate structural features that regulate selective dehydrogenation rather than oxygenation. These goals will be realized through the following aims: 1) To determine the structures of the reactive intermediates that are produced by dehydrogenation of prototypical substrates, and characterize enzyme preferences for dehydrogenation vs. oxygenation of the substrates; 2) To characterize the mechanisms of inactivation of each P450 enzyme by its specific inactivator; 3) To define the active-site parameters that control the mechanisms of dehydrogenation and bioactivation of toxicants by P450 enzymes; and 4) To use the dehydrogenation substrates that covalently modify the P450 apoproteins to elucidate critical active-site residues that direct the dehydrogenation mechanism, or that control inhibitor/substrate access channels, binding, or product release. The enzyme/substrate pairs are CYP2F3/3-methylindole, CYP3A4/zafirlukast, CYP2El/capsaicin, and CYP2B6/tamoxifen. The long-term goals of this research are to elucidate the mechanisms of cytochrome P450-mediated dehydrogenation of xenobiotics in processes that generate toxic electrophilic intermediates, to assess the potential harm engendered by these toxic intermediates to human health, and to utilize mechanistic information to predict dehydrogenation, and concomitant toxicities and/or enzyme inactivation (altered drug metabolism), of new drugs and xenobiotics.

描述（由申请人提供）：通过细胞色素P450氧合机制将异种生物学生物活化至有毒中间体是一个众所周知的过程。然而，几种P450酶（例如1A2、2B6、2E1、2F1、3A4和4B1）通过脱氢途径的产生，仅仅研究了选择性脱氢而不是氧化的机制。几种脱氢中间体具有反应性，以至于它们通常通过活性位点亲核残基的烷基化使P450酶失活。关于这些特定P450酶的催化行为及其对脱氢而不是氧化底物的倾向的研究非常需要。这项研究的假设是：促进电子转运的独特催化机制，该促进的电子转运决定了某些P450酶的脱氢作用会导致异种生物介导的损伤和人类药物代谢改变。本应用的具体目标是确定特定细胞色素p450酶的酶有源位点环境的特征，并定义调节选择性脱氢而不是氧化的底物结构特征。这些目标将通过以下目的实现：1）确定由原型底物脱氢产生的反应性中间体的结构，并表征酶偏好的酶偏好，以脱氢与底物的氧化； 2）表征每种P450酶通过其特异性灭活剂的灭活机制； 3）定义控制P450酶对有毒物质脱氢机制和生物活化机制的活性位点参数； 4）使用共价修改P450载蛋白的脱氢底物来阐明指导脱氢机制的关键活性位点残基，或者控制抑制剂/底物访问通道，结合或产品释放。酶/底物对是CYP2F3/3-甲基吲哚，CYP3A4/Zafirlukast，CYP2EL/辣椒素和CYP2B6/Tamoxifen。这项研究的长期目标是阐明细胞色素p450介导的异种生物学的脱氢作用的机制，这些过程在产生有毒的亲电中间体的过程中，评估这些有毒中间体对人类健康产生的潜在伤害，并利用机械性信息来预测机械性（Orniz Inter）（抗性药物和综合性毒性），或者是与综合性毒性的毒性，或者代谢），新药和异种生物。