Regulation of NADH: ubiquinone oxidoreductase (complex *

NADH 的调节：泛醌氧化还原酶（复合物 *

• 批准号: 6548756
• 负责人: Gary Cecchini
• 金额: $ 4.17万
• 依托单位: NORTHERN CALIFORNIA INSTITUTE/RES/EDU
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-08-15 至 2005-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6548756
• 关键词: Commonwealth of Independent States; NAD(P)H oxidoreductase; SDS polyacrylamide gel electrophoresis; active sites; binding sites; complementary DNA; cooperative study; enzyme structure; enzyme substrate; laboratory rat; ligands; mitochondria; mutant; myocardial ischemia /hypoxia; nucleic acid sequence; protein purification; reperfusion; ubiquinone

DESCRIPTION (provided by applicant) NADH:ubiquinone oxidoreductase (complex I) is an energy-transducing membrane-bound protein complex that serves as a major entry point for electrons to the mitochondrial respiratory chain. The multisubunit enzyme complex is encoded by 43 separate genes in mammals. The mammalian enzyme shows extremely complex kinetic behavior, in all forms that retain natural electron transfer pathways. The complex kinetic behavior suggests that two distinct slowly equilibrating forms of the enzyme exist. The active (A) form is catalytically capable to reduce ubiquinone, whereas, the de-active (D) form is not able to transfer electrons to ubiquinone in the membrane. The equilibrium between these two forms is under a refined control by a number of regulatory factors (i.e., pH, divalent cations, temperature, the ubiquinone/ubisemiquinone/ubiquinol ratio). This research will primarily be done in Russia as an extension of NIH grant R01 GM61606. The proposed collaborative work is to extend studies of the regulatory characteristics of the enzyme at the molecular mechanistic level by using a combination of biochemical and molecular approaches. The studies will be done in physiologically relevant systems such as intact hearts and mitochondria, as well as, the kinetic and biochemical approaches in the foreign Co-P.l.'s laboratory. Two major aspects of the ND transition are to be investigated. One is to elucidate the molecular mechanisms of the slow interconversion between the A and D forms involved in the red/ox-dependent D to A transition; and the extremely temperature-dependent A to D transition. Natural ligands and modifiers which affect the rate and equilibrium of the ND transition in vitro will be identified. The second aim focusses on the complex I A/D transition in intact mitochondria in relation to the experimental model of ischemia/reperfusion in Langendorff-pertused rat hearts. The focus of these studies is to determine the possible physiological importance of the A/D transition of complex I as it responds to alterations in cellular metabolism. The catalytic activity of complex I is important for cellular physiology and a number of diseases (diabetes, cardiomyopathy) have been associated with defects in complex I. The research to be undertaken will shed light on the modulation of enzyme activity of this important respiratory complex.

描述(申请人提供)NADH：泛醌氧化还原酶(复合体I)是一种能量转导的膜结合蛋白复合体，是电子进入线粒体呼吸链的主要入口点。哺乳动物的多亚单位酶复合体由43个不同的基因编码。哺乳动物的酶表现出极其复杂的动力学行为，所有形式都保留了自然的电子转移途径。复杂的动力学行为表明，该酶存在两种不同的缓慢平衡形式。活性(A)形式能够催化还原泛醌，而非活性(D)形式不能将电子转移到膜中的泛醌。这两种形式之间的平衡受到一系列调节因素(即pH、二价阳离子、温度、泛醌/泛喹酮/泛喹酚比率)的精细控制。这项研究将主要在俄罗斯进行，作为NIH拨款R01 GM61606的延伸。拟议的合作工作是通过结合使用生物化学和分子方法，在分子机制水平上扩展对酶调节特性的研究。这些研究将在与生理相关的系统中进行，如完整的心脏和线粒体，以及国外公司S实验室的动力学和生化方法。ND过渡的两个主要方面有待研究。一是阐明依赖于RED/OX的D-A转变和极端依赖温度的A-D转变所涉及的A和D形式之间缓慢相互转化的分子机制。将确定在体外影响ND转换速率和平衡的天然配体和修饰剂。第二个目标集中在完整线粒体的复杂的A/D转变与朗宁多夫灌流的大鼠心脏缺血/再灌注实验模型的关系上。这些研究的重点是确定复合体I的A/D转换在它对细胞代谢变化的反应中可能的生理重要性。复合体I的催化活性对细胞生理学很重要，许多疾病(糖尿病、心肌病)都与复合体I的缺陷有关。即将进行的研究将有助于阐明这种重要的呼吸复合体的酶活性调节。