ISCHEMIC/REPERFUSION DAMAGE TO THE RIESKE IRON PROTEIN

缺血和再灌流期间心肌损伤增加 年迈的心脏。当时在复合体III中存在两个串联缺陷 老化心脏的再灌流：Rieske铁-硫的损伤 蛋白质(Isp)和细胞色素c结合位点上与衰老相关的缺陷。 我们的初步证据表明，该互联网服务供应商是 与线粒体功能缺陷相关的损伤 缺血症。开发了一种分级方法来识别该化学物质 Isp在再灌流期间遭受的氧化损伤的性质， 可能是活性物质的化学性质导致了 蛋白质修饰，以及观察到的修饰的相关性 再灌注性心肌损伤。四个具体目标是 被描述为实现这些目标。第一是发展和发展 展示高效的生化和质谱学方法 氧化损伤的结构鉴定。第二个具体目标 研究氧化损伤的光谱，这可能会对 Isp的可溶性催化结构域，简称铁硫 片断，然后在复合体内的ISP集成层面上 第三个具体目标是研究这些氧化剂如何改变功能 当isp在完整的线粒体内运作时。这一具体目标 允许对潜在的预期产品进行表征 与生理相关的氧化剂和脂类生成的活性物种 过氧化。将在下列情况下检查氧化修饰的模式 电子流量随特征良好的缓蚀剂而变化。 关于电子传输的问题。第四个具体目标是确定 在缺血-再灌流过程中产生的氧化修饰。对.的使用 /180/2将允许检测到的氧化修饰 在再灌流后分离的isp中存在明确的相关性 与再灌流损伤有关。鉴定出的蛋白质的比较 使用前3个目标积累的数据进行修改将 确定再灌流可能产生的活性物质，以及 建议和线粒体为其产生和线粒体提供一个位置 蛋白质损伤的机制。这些信息将在第四章中使用 目的设计和评价以机械为基础的治疗干预措施 改善老龄心脏缺血再灌流损伤。

Myocardial damage during ischemia and reperfusion is increased in the elderly heart. Two tandem defects are present in complex III at the time of reperfusion in the aging heart: damage to the Rieske iron-sulfur protein (ISP) and a aging-related defect in the cytochrome c binding site. Our preliminary evidence has identified the ISP as a prominent site of damage associated with mitochondrial functional deficits following ischemia. A hierarchical approach is developed to identify the chemical nature of oxidative damage suffered by the ISP during reperfusion, the probably chemical nature of the reactive species responsible for the protein modification, and the correlation of the observed modification with reperfusion induced myocardial injury. Four specific aims are described to accomplish these goals. The first is to develop and demonstrate biochemical and mass spectrometric methods for efficient structural identification of oxidative-damage. The second specific aim investigates the spectrum of oxidative damage that can be inflicted on the soluble catalytic domain of the ISP, referred to as the iron-sulfur fragment, and then at the level of integration of the ISP within complex III. The third specific aim examines how these oxidants alter the function of the ISP when it functions within intact mitochondria. This specific aim permits the characterization of the anticipated products from potentially physiologically relevant oxidants and reactive species generated by lipid peroxidation. The pattern of oxidative modifications will be examined when the electron flow is varied in response to well characterized inhibitors of electron transport. The fourth specific aim is to determine the oxidative modifications arising during ischemia-reperfusion. The use of /180/2 during reperfusion will permit the oxidative modifications detected in the ISP isolated following reperfusion to be unequivocally correlated with the reperfusion injury. Comparison of the identified protein modifications with the data accumulated from the first 3 aims will identify the probably reactive species generated by reperfusion, and suggest and mitochondrial a mitochondrial site for its generation and mechanism of protein damage. This information will be used in the fourth aim to design and evaluate mechanistic-based therapeutic interventions to ameliorate damage during ischemia and reperfusion in the aging heart.