STUDIES OF TRIMETHYLAMINE DEHYDROGENASE

三甲胺脱氢酶的研究

• 批准号: 6180830
• 负责人: Russ Hille
• 金额: $ 15.59万
• 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/6180830
• 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）该项目的总体目标 拟议的研究旨在更深入地了解其作用机制 三甲胺脱氢酶 (TMADH)，一组重要酶的成员 含有多个氧化还原活性中心并具有整体催化作用 涉及这些位点之间电子转移的机制。此外 它们的生理意义，这些蛋白质非常有用 用于检查控制磁力和磁力的因素的系统 这些中心之间的电子转移相互作用。催化反应为 TMADH，三甲胺氧化脱甲基为二甲胺 甲醛，对于了解甲醛具有相当大的内在兴趣 生物系统中异咯嗪环的化学性质。这 化学与许多其他酶相关，包括临床上的酶 重要的酶，如单胺氧化酶。 拟议的工作以 PI 之前的工作为基础，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 反应表征 使用时间分辨晶体学；形容异常坚强的人 TMADH 的黄素和 Fe4S4 半醌中心之间的磁相互作用； 电化学和电子传输特性的检查 三甲胺脱氢酶；以及三甲胺之间相互作用的表征 脱氢酶及其生理电子受体，和 电子传递黄素蛋白。以三维结构为导向 三甲胺脱氢酶的特定氨基酸残基的作用 对于这些区域中的每一个区域，酶的活性位点将是 通过定点诱变来解决。这项工作的目的是 构建结构与功能关系的全面图景 三甲胺脱氢酶，进一步加深了我们对三甲胺脱氢酶的基本了解 该酶和相关酶的作用机制。