Myoglobin as a Nitrite Reductase that Regulates Hypoxic Cardiac NO Signaling

描述（由申请人提供）：人们逐渐认识到，血红素蛋白（如肌红蛋白和血红蛋白）催化一氧化氮（NO）和亚硝酸盐的代谢，从而调节细胞和组织对缺氧的反应。在过去的五年里，我们的小组已经发表了生化和生理研究，表明血红蛋白作为亚硝酸盐还原酶的一种新功能，在生理和病理缺氧条件下产生NO，这表明肌红蛋白，另一种主要的血红素珠蛋白，也可能具有介导氮氧化物代谢的能力，受氧张力的影响。类似于细菌的亚硝酸盐还原酶，协同质子和电子转移反应，亚硝酸盐减少阴离子NO。我们还发现了一种新的亚硝酸盐脱水酶活性，将两个亚硝酸盐分子转化为高度扩散的亚硝化分子N2 O3，使有效的NO信号在血红素丰富的环境。在目前的研究计划中，NO/N2 O3信号通路将在肌红蛋白中探索，特别关注通过a）通过调节线粒体电子转移反应调节细胞代谢和B）激活缺血-再灌注损伤后的净细胞保护性细胞信号反应来调节细胞对缺氧和缺血应激的反应的能力。这些概念将通过测试肌红蛋白介导的亚硝酸盐代谢调节缺氧NO信号传导和促进NO在心脏中的生物利用度的假设来解决。更具体地说，在体外生物化学和细胞培养系统以及心肌梗死的体内模型中使用突变肌红蛋白，我们的目的是1）确定缺氧时肌红蛋白亚硝酸盐还原酶活性的分子和酶机制，2）定义肌红蛋白衍生的NO和N2 O3的关键分子靶点，和3）确定肌红蛋白在缺血/再灌注后调节下游细胞保护信号（细胞凋亡和线粒体生物发生）中的作用。成功完成拟议的研究计划将促进我们对肌红蛋白的生物学功能和亚硝酸盐在心血管系统中的生理和药理学潜力的理解。 公共卫生相关性：心脏病发作后，心脏中的部分组织死亡。一种名为肌红蛋白的心脏蛋白质可以将饮食中发现的亚硝酸盐转化为一氧化氮，一氧化氮是一种保护心脏细胞免于死亡的化学物质。该项目将研究亚硝酸盐和肌红蛋白如何共同作用，以便在心脏病发作期间使用亚硝酸盐来保护心脏。

DESCRIPTION (provided by applicant): There is an emerging appreciation that heme proteins such as myoglobin and hemoglobin catalyze the metabolism of nitric oxide (NO) and nitrite, and thereby modulate cell and tissue responses to hypoxia. Over the last five years our group has published biochemical and physiological studies that suggest a novel function for hemoglobin as a nitrite reductase that generates NO under physiological and pathological hypoxia, suggesting that myoglobin, the other major heme globin, may also possess a capability to mediate the metabolism of oxides of nitrogen in a manner influenced by oxygen tension. Analogous to the bacterial nitrite reductases, a concerted proton and electron transfer reaction to nitrite reduces the anion to NO. We have also discovered a novel nitrite anhydrase activity that converts two nitrite molecules into the highly diffusible, nitrosating molecule N2O3, allowing efficient NO signaling in a heme rich environment. In the current research proposal, the NO/ N2O3 signaling pathways will be explored in myoglobin, with special focus on an ability to regulate cellular responses to hypoxic and ischemic stress by a) modulating cellular metabolism via the regulation of mitochondrial electron transfer reactions and b) activating net cytoprotective cell signaling reactions after ischemia-reperfusion injury. These concepts will be addressed by testing the hypothesis that myoglobin-mediated nitrite metabolism regulates hypoxic NO signaling and promotes NO bioavailability in the heart. More specifically, using mutant myoglobin proteins in in vitro biochemical and cell culture systems as well as an in vivo model of myocardial infarction, we aim 1) to determine the molecular and enzymatic mechanisms underlying the nitrite reductase activity of myoglobin in hypoxia, 2) define critical molecular targets of myoglobin-derived NO and N2O3, and 3) to determine the role of myoglobin in regulating downstream cytoprotective signaling (apoptosis and mitochondrial biogenesis) after ischemia/reperfusion. Successful completion of the proposed research plan will advance our understanding of the biological function of myoglobin and the physiological and pharmacological potential of nitrite in the cardiovascular system. PUBLIC HEALTH RELEVANCE: After a heart attack, part of the tissue in the heart dies. A heart protein called myoglobin can convert nitrite, a chemical found in the diet, to nitric oxide, a chemical that protects heart cells from death. This project will investigate how nitrite and myoglobin work together so that nitrite can be used to protect the heart during a heart attack.