MECHANISMS THAT REGULATE ENZYMES THAT METABOLIZE CIS UNSATURATED FATTY ACIDS

工作总结：目标是了解花生四烯酸和亚油酸代谢的生化机制。对PGHS-1的研究表明，酪氨酸基的形成可能是引发花生四烯酸氧化的反应中间体。最近，我们发现NO与酪氨酸基发生反应，使之转化为一种新的自由基，经ESR分析表征为亚氨氧基。亚氨氧基衰变为硝基酪氨酸，导致PHS上的酪氨酸部分硝化。此外，其生成的硝基酪氨酸经常被用作氧化应激的指示物。通过NO与酪氨酸基的反应，我们确定了PHS中特定的酪氨酸部分为酪氨酸-385。这表明酪氨酸-385在催化过程中被氧化为酪氨酸基，并表明该残基可能是活性部位或酶失活部位。未来的研究将集中在了解COX酪氨酸自由基与NO的相互作用及其对COX活性的影响。此外，我们目前正在测试一种假设，即硝基酪氨酸的测量主要是通过代谢活性的COX-2和iNOS形成的NO的相互作用发生的。了解调节这些酶的活性和表达的生化机制将为潜在地控制或预防这些疾病状况提供新的见解。-前列腺素-H-合成酶、COX-1、COX-2、酪氨酸基

Summary of Work: The goal is to develop an understanding of the biochemical mechanisms responsible for the metabolism of arachidonic acid and linoleic acid. Studies with PGHS-1 indicate that a tyrosyl radical is formed which could be the reactive intermediate that initiates arachidonic acid oxygenation. Recently, we have shown that NO reacts with the tyrosyl radical converting the radical to a new radical characterized by ESR analysis as an iminoxyl radical. The iminoxyl radical decays to nitrotyrosine resulting in the nitration of the tyrosine moiety on PHS. In addition, its formation of nitrotyrosine is often used as an indicator of oxidative stress. We have identified the specific tyrosine moiety in PHS nitrated by the reaction of NO with tyrosyl radical as Tyrosine-385. This indicates that tyrosine-385 is oxidized to tyrosyl radical during catalysis and suggests this residue is a likely active site or is the site of enzyme inactivation. Future studies will focus on understanding the interaction of Cox tyrosine radical with NO and its effect on Cox activity. In addition, we are currently testing the hypothesis that measurement of nitrotyrosine, a marked for oxidative stress, occur mainly by the interaction of metabolically active Cox-2 and NO formed by iNOS. Understanding the biochemical mechanisms which regulate the activities and expression of these enzymes will provide new insights into potentially controlling or preventing these disease conditions. - Prostaglandin-H-synthase, COX-1, COX-2, tyrosyl radical