INACTIVATORS OF LIPOXYGENASE

脂氧合酶灭活剂

• 批准号: 3292581
• 负责人: CHARLES H CLAPP
• 金额: $ 5.96万
• 依托单位: BUCKNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1992-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3292581
• 关键词: chemical structure function; chemical synthesis; deuterium; enzyme inhibitors; enzyme mechanism; iodine; lipid peroxides; lipoxygenase; molecular site; oleate; radiotracer; stoichiometry; thiols

Soybean lipoxygenase is a nonheme iron protein (MW = 94,500) that catalyzes the oxygenation of linoleic acid to 13-hydroperoxy-9,11- octadecadienoic acid (13-HPOD). The goals of this project are to understand the chemical properties and catalytic mechanism of this enzyme and to develop strategies that can be applied to the design of inhibitors of several physiologically and pharmaceutically interesting mammalian lipoxygenases. 12-Iodo-cis-9-octadecenoic acid (12-IODE) is a time-dependent, irreversible inactivator of soybean lipoxygenase. Inactivation requires both 13-HPOD (or some other lipid hydroperoxide) and 02. These observations suggest that 12-IODE is a suicide inactivator, and this idea has been strengthened by our finding that inactivation is accompanied by formation of about 9 iodide ions per molecule of enzyme inactivated. 9,11-Octadecadienoic acid is formed in quantities that are less than stoichiometric with iodide formation, which implies that other organic products are formed from 12-IODE. 02 uptake has also been detected. During the period covered by this application we will use 1-14C-12-IODE to identify the remaining organic products from the action of lipoxygenase on 12-IODE. We will also use this material to determine whether the carbon skeleton of 12-IODE becomes incorporated into the inactivated enzyme, and of so, we will carry out degradation studies on the labelled enzyme to determine how the inhibitor has become attached. In addition, we will synthesize 11,11-dideuterio- 12-IODE and use this material to test our working hypothesis by measuring the effect of deuterium substitution on the rate of iodide formation and on the rate and extent of inactivation. We have also found that 12-mercapto-cis-9-octadecenoic acid (12- HSODE) is a time-dependent, irreversible inactivator of lipoxygenase. Inactivation is accompanied by consumption of about 67 molecules of 12 HSODE per molecule of enzyme inactivated. These observations suggest that 12-HSODE is also a suicide inactivator. During the period covered by this application we will identify the products produced by the action of lipoxygenase on 12-HSODE, attempt to determine how the inactivated enzyme has been modified, and carry out other experiments to elucidate the inactivation mechanism.

大豆脂氧酶是一种非血红素铁蛋白（MW = 94,500） 催化亚油酸将亚油酸的氧合催化为13-氢氧化物-9,11- Octadecadecadienoic酸（13-HPOD）。 该项目的目标是 了解这一点的化学特性和催化机制 酶并制定可应用于设计的策略 几种生理和药物的抑制剂 有趣的哺乳动物脂氧酶。 12- iodo-cis-9-octadecenoic Acid（12-iode）是时间依赖的， 大豆脂氧酶的不可逆转灭活剂。 失活 需要13-HPOD（或其他一些脂质氢过氧化物）和02。 这些观察结果表明12-IODE是自杀灭活剂， 我们的发现加强了这个想法 灭活伴随着大约9个碘化离子 酶的分子灭活。 9,11-otctadecadecadecadienoic Acid是 与碘化物的化学计量学的数量相比形成 形成，这意味着形成了其他有机产品 从12个IODE。 还检测到02吸收。 在此期间 在此应用程序中，我们将使用1-14C-12-IODE识别 脂氧合酶对作用的剩余有机产品 12-iode。 我们还将使用此材料来确定是否 12-iode的碳骨架被纳入 灭活酶，因此，我们将进行降解 对标记酶的研究，以确定抑制剂的方式 变得依恋。 此外，我们将合成11,11-dideuterio- 12-iode并使用此材料来检验我们的工作假设 测量氘取代对 碘化物形成和灭活率和程度。 我们还发现，12-甲基CIS-CIS-9-二十二烯酸（12-- hsode）是时间依赖的，不可逆的灭活因子 脂氧合酶。 失活伴随着大约的消费 每个分子灭活的酶的12 Hsode的67个分子。 这些 观察结果表明12-Hsode也是自杀灭活剂。 在此应用程序所涵盖的期间，我们将确定 Lipoxygonase对12-Hsode的作用产生的产品， 试图确定如何修饰灭活酶， 并进行其他实验以阐明失活 机制。