MEMBRANE BOUND DETOXICATION ENZYMES

潜在毒性异源物质的代谢与解毒 分子依赖于几个膜结合分子的参与 包括细胞色素P-450、微体环氧化物在内的酶 水解酶和UDP-葡萄糖醛酸基转移酶。环氧化物水解酶 催化水与环氧化物的反式加成反应 由细胞色素P-450作用于不饱和 碳氢化合物。UDP-葡萄糖醛酸基转移酶催化转运蛋白 从UDP-葡萄糖醛酸基到疏水分子的克鲁醛酸基 带有亲核官能团，如-OH、-SH、-NH2和- 太棒了。这些反应的最终结果是提高了溶解度， 母体分子的运输和排泄。具体目标是 拟议的项目期主要集中在以下三个一般领域 查询包括对表达系统的改进 蛋白质，酶的作用机制的阐明 并用X射线对酶的结构进行了初步研究 结晶学和生化技术。《公约》的具体目标 关于微粒体环氧化物水解酶的计划是：(I) 用预稳态法和稳态法阐明动力学机理 国家技术；(2)化学机理的定义 酯类化合物的定点诱变催化作用及其表征 中间体；(3)潜在的基于机理的抑制剂的评价 以及(Iv)生产X射线衍射级质量的单晶 水晶。UDP调查的具体目的是- 葡萄糖醛酸基转移酶包括：(I)表达的优化 杆状病毒或细菌系统中的苯酚UDPGT； (Ii)对酶的构象特异性的探索 二氢二醇底物。这些研究预计将增加我们的 对新陈代谢分子细节的理解 环境污染物和毒品，最终提高我们的能力 来预测新的潜在有毒化合物的代谢情景。

The metabolism and detoxication of potentially toxic xenobiotic molecules relies on the involvement of several membrane-bound enzymes including the cytochromes P-450, microsomal epoxide hydrolase and UDP-glucuronosyltransferase. Epoxide hydrolase catalyzes the trans-addition of water to epoxides that are often generated by the action of the cytochromes P-450 on unsaturated hydrocarbons. UDP-glucuronosyltransferase catalyzes the transfer of the clucuronyl group from UDP-glucuronate to hydrophobic molecules bearing nucleophilic functional groups such as -OH, -SH, -NH2 and - COOH. The net result of these reactions is to enhance the solubility, transport and excretion of the parent molecules. The specific aims for the proposed project period are focused on three general areas of inquiry including the improvement of expression systems for the proteins, the elucidation of the mechanisms of action of the enzymes and preliminary investigations of the structure of the enzymes by X-ray crystallography and biochemical techniques. The specific aims of the project with respect to microsomal epoxide hydrolase are: (I) the elucidation of the kinetic mechanism by pre-steady-state and steady- state techniques; (ii) the definition of the chemical mechanism of catalysis by site-directed mutagenesis and characterization of the ester intermedicate; (iii) evaluation of potential mechanism-based inhibitors of the enzyme; and (iv) production of X-ray diffraction-quality single crystals. The specific aim for the investigations of UDP- glucuronosyltransferase include: (I) optimization of the expression of the phenol UDPGT in a baculovirus-based or bacterial systems and ; (ii) an exploration of the conformer specificity of the enzyme toward dihydrodiol substrates. These studies are anticipated to increase our understanding of the molecular details of the metabolism of environmental pollutants and drugs and eventually enhance our ability to predict metabolic scenarios for new, potentially toxic compounds.