Biomimetic Studies of NO-binding Respiratory Chain Hemes

描述（由申请人提供）：呼吸作用是在细胞水平上消耗双氧，为代谢过程提供能量，由终端氧化酶如细胞色素c氧化酶（CcO）介导。最近的实验工作表明，CcO对能源生产的调节比迄今所认识到的要复杂得多。一个特殊的因素是，人们认识到一氧化氮（NO）是一种气态自由基，在体内作为二氧结合CcO的竞争性抑制剂具有重要作用。除了正常的，尽管人们对NO在体内的调节作用知之甚少，这一重要发现开辟了一个前景，即相当多的病理状况是由NO的异常水平及其随后与蛋白质的反应引起的，如电子传递链（其中CcO是末端元件）。帕金森氏症、阿尔茨海默氏症、亨廷顿氏症和ALS都与NO/CcO这对夫妇有关。虽然有大量的现象学实验证明了能量调节与NO水平之间的联系，但NO的反应机制以及CcO血红素/铜活性位点中间体的物理化学特征与调节和病理的关系仍然存在争议。值得注意的是，CcO的祖先一氧化氮还原酶（NOR）不受NO的抑制，而是通过释放能量将其还原为N20，类似于CcO将O2还原为H2O的反应。一氧化氮减少一氧化氮的机制及其结构与这种活性的关系尚不清楚。为了继续了解末端氧化酶活性位点小分子的仿生反应，本项目旨在表征NO与血红素/铜（CcO）和血红素/铁（NOR）仿生复合物的反应。通过对先前开发的配体系统的光谱和电化学研究，使用许多双金属配合物与金属离子组合，不可能获得生物学上的，但对于分离体内酶特性的因素至关重要，我们的目标是回答关于NO与CcO和NOR反应差异的许多问题。反应中间体的鉴定，它们的反应性，泄漏和可能的生物后果将是研究的重点。重点将放在动力学和机理研究，以阐明CcO和NOR的推定反应途径。

DESCRIPTION (provided by applicant): Respiration, the consumption of dioxygen at the cellular level to provide energy for metabolic processes, is mediated by terminal oxidases such as cytochrome c oxidase (CcO). Recent experimental work has revealed more complexity in the regulation of energy production by CcO than hitherto appreciated. One specific factor has been the realization that nitric oxide (NO), a gaseous free radical, has an important role in vivo as a competitive inhibitor of dioxygen binding to CcO. Aside from the normal, although poorly understood, regulatory role of NO in vivo this important discovery opened the prospect that a considerable number of pathological conditions arise from abnormal levels of NO and its subsequent reactions with proteins such as those of the electron transport chain (of which CcO is terminal element). Parkinson's, Alzheimer's, Huntington's disease and ALS have been linked to the NO/CcO couple. While there are a plethora of phenomenological experiments demonstrating the connection between energy regulation and NO levels, the mechanism of reaction of NO and the physicochemical characteristics of intermediates at the heme/copper active site of CcO as they relate to regulation and pathology remain controversial. Significantly, an ancestral progenitor of CcO, Nitric Oxide Reductase (NOR), is not inhibited by NO, but reduces it to N20 with release of energy, a reaction analogous to the reduction of O2 to H2O by CcO. The mechanism by which NOR reduces NO and the relationship of its structure to this activity is little known. Continuing a long-term goal of understanding biomimetic reactions of small molecules at the active sites of terminal oxidases, this project aims to characterize the reaction of NO with heme/copper (CcO) and heme/iron (NOR) biomimetic complexes. Through spectroscopic and electrochemical investigation of previously-developed ligand systems, using numerous bimetallic complexes with metal ion combinations not possible to obtain biologically, but essential for isolating the factors that are contributing to the enzymatic characteristics in vivo, we aim to answer many questions concerning the differences in NO reaction with CcO and NOR. The identification of reaction intermediates, their reactivity, leakage and possible biological consequences will be a focus for study. An emphasis will be placed on kinetic and mechanistic studies to elucidate the putitive reaction pathways of CcO and NOR.