SPECTROELECTROCHEMICAL STUDY OF SELECTED FLAVOPROTEINS

所选黄素蛋白的光谱电化学研究

• 批准号: 3276909
• 负责人: MARIAN T. STANKOVICH
• 金额: $ 13.7万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1981
• 资助国家: 美国
• 起止时间: 1981-07-01 至 1993-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3276909
• 关键词: acidity /alkalinity; acyl coA dehydrogenases; bacteria; electrical potential; electrochemistry; electron transport; enzyme inhibitors; enzyme mechanism; enzyme structure; enzyme substrate; flavoproteins; hydroxybenzoate; ligands; mitochondria; oxidation reduction reaction; oxidoreductase; oxygenases; spectrometry; swine; thermodynamics

Flavoproteins catalyze many biologically important electron- transfer reactions. We have developed and used a spectroelectrochemical method to systematically study the thermodynamic properties of two flavoprotein groups, the oxidases and the dehydrogenases. We plan to finish the characterization of the three main groups of flavoproteins by studying the monooxygenases, and we propose to extensively study the ligand binding properties of the flavoprotein dehydrogenase enzymes involved in the beta-oxidation of fatty acids. A parallel study will be performed on ligand binding to the monooxygenases. The beta-oxidation of fatty acids is an important energy conversion process in heart muscle, skeletal muscle, liver and kidney. It is the primary energy source for heart muscle, providing 90% of this energy. The beta-oxidation process is regulated by the acyl-CoA dehydrogenases (ACD), which catalyze the first step. It has been postulated that the ACD's are isopotential with their substrates and that product binding controls the completeness of the reaction. Using enzymes from a bacterial source we have shown that the substrate is not isopotential with the enzyme and that the binding of the substrate or product to the reduced form has shifted the potential significantly positive. A similar shift would be of more significance in a mammalian system. Work done by others suggests that such shifts are occurring. This work taken together strongly suggests that for the mammalian system, substrate-bound ACD is isopotential with electron transferring flavoprotein (ETF) and with ETF-ubiquinone oxidoreductase. This suggestion has profound implications on the energetics of the beta-oxidation process. This work suggests that a further redox study will give new inisights into regulation (and inhibition) of oxidation. Our aims are to measure the electron energy levels (redox potentials) of free enzymes and substrates, then to measure the effect of ligand binding on those levels. Thus, we will be able to assess the relative importance of binding in controlling catalytic reactions. Electrochemistry is the best method to examine these interactions because we can clearly show that electron-energy levels change in a measurable way upon ligand binding. We will perform a comparative study of acyl-CoA dehydrogenases, from mitochondria, bacteria which are mitochondrial precursors and anaerobic bacteria bound to substrates, products, inhibitors, and their respective ETF's.

黄素蛋白催化许多生物学上重要的电子- 转移反应 我们开发并使用了 光谱电化学方法系统地研究了 热力学性质的两个黄素蛋白组， 氧化酶和脱氢酶。 我们计划完成 通过以下方法表征三种主要的黄素蛋白组： 研究单加氧酶，我们建议广泛研究 黄素蛋白脱氢酶的配体结合特性 参与脂肪酸β-氧化的酶。 并行 将对配体与单加氧酶的结合进行研究。 脂肪酸的β-氧化是一种重要的能量 心肌、骨骼肌、肝脏和 肾 它是心肌的主要能量来源， 提供90%的能量 β-氧化过程是 由酰基辅酶A脱氢酶（ACD）调节，其催化 第一步。 据推测，ACD是 与其底物和产物结合等电位 控制反应的完全性。 使用来自 细菌来源，我们已经表明，底物不是 与酶的等电位和底物的结合 或产品的还原形式已经改变了潜在的 非常积极。 类似的转变将更多 在哺乳动物系统中的重要性。 其他人所做的工作 表明这种转变正在发生。 这项工作结合在一起 强烈表明，对于哺乳动物系统，底物结合 ACD与电子转移黄素蛋白（ETF）等电位 和ETF-ubiquinone氧化还原酶。 这一建议 对β-氧化的能量学有着深远的影响 过程 这项工作表明，进一步的氧化还原研究将使 对氧化调节（和抑制）的新见解。 我们 目的是测量电子能级（氧化还原电位） 游离酶和底物，然后测量 配体结合的水平。 这样，我们就可以评估 结合在控制催化反应中的相对重要性。 电化学是检测这些的最好方法 因为我们可以清楚地表明， 水平在配体结合时以可测量的方式变化。 我们将 进行酰基辅酶A脱氢酶的比较研究， 线粒体，细菌是线粒体前体， 与底物、产物、抑制剂结合的厌氧菌， 各自的ETF。