STUDIES OF TRIMETHYLAMINE DEHYDROGENASE

三甲胺脱氢酶的研究

• 批准号: 2908202
• 负责人: Russ Hille
• 金额: $ 17.14万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-01 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2908202
• 关键词: X ray crystallography; aminoacid; electron spin resonance spectroscopy; electron transport; enzyme activity; enzyme mechanism; flavins; intermolecular interaction; metalloenzyme; methylation; oxidizing agents; oxidoreductase; protein structure function; site directed mutagenesis; tertiary amine

DESCRIPTION: (adapted from applicant's abstract) The overall goal of the proposed research is to gain deeper insight into the mechanism of action of trimethylamine dehydrogenase (TMADH), a member of an important group of enzymes that contain multiple redox-active centers and which have overall catalytic mechanisms involving electron transfer between these sites. In addition to their physiological significance, these proteins serve as extremely useful systems in which to examine the factors which govern the magnetic and electron-transfer interactions between such centers. The reaction catalyzed by TMADH, the oxidative demethylation of trimethylamine to dimethylamine and formaldehyde, is of considerable intrinsic interest in understanding the chemistry available to the isoalloxazine ring in biological systems. This chemistry is relevant to a number of other enzymes, including such clinically important enzymes as monoamine oxidase. The proposed work builds on previous work by the PI and has the following principal objectives: Examination of the role of specific amino acid residues in the reaction catalyzed by TMADH; characterization of the reaction of TMADH using time-resolved crystallography; characterization of the unusually strong magnetic interaction between the flavin and Fe4S4 semiquinone centers of TMADH; examination of the electrochemical and electron-transferring properties of TMADH; and characterization of the interaction between trimethylamine dehydrogenase and its physiological electron acceptor, and electron-transferring flavoprotein. Guided by the three-dimensional structure of trimethylamine dehydrogenase, the roles of specific amino acid residues in the active site of the enzyme with regard to each of these areas will be addressed by site-directed mutagenesis. The intention of this work is to construct a comprehensive picture of the relationship of structure to function in trimethylamine dehydrogenase that furthers our basic understanding of the mechanism of this and related enzymes.

描述：（改编自申请人的摘要） 拟议的研究是为了深入了解 三甲胺脱氢酶（TMADH），一组重要酶的成员 其中包含多个氧化还原活性中心，并且具有总体催化 这些位点之间涉及电子转移的机制。此外 它们的生理意义，这些蛋白质非常有用 检查控制磁性因素的系统 这些中心之间的电子转移相互作用。反应由 tmadh，三甲胺对二甲胺和二甲胺的氧化脱甲基化和 甲醛，在理解 在生物系统中可用于异丙沙嗪环的化学。这 化学与许多其他酶有关，包括这种临床 重要酶作为单胺氧化酶。 拟议的工作是基于PI以前的工作，并具有以下内容 主要目标：检查特定氨基酸残基的作用 在tmadh催化的反应中； tmadh反应的表征 使用时间分辨的晶体学；异常强壮的表征 黄素和Fe4s4 semiquinone中心之间的磁相互作用； 检查电化学和电子转移特性 tmadh;和三甲胺之间的相互作用的表征 脱氢酶及其生理电子受体，以及 电子转移黄蛋白。在三维结构的指导下 三甲胺脱氢酶，特异性氨基酸残基在 关于这些区域的每个区域的酶的活跃部位将是 通过定向诱变来解决。这项工作的目的是 构建结构与功能关系的全面图片 在三甲胺脱氢酶中，我们对 该酶的机制。