FUNCTIONAL STUDIES OF HUMAN AND SOYBEAN LIPOXYGENASE

描述：脂氧合酶(LOS)是一种催化 含脂肪的1，4-顺式、顺式戊二烯中掺入氧气的研究 酸，广泛分布于动植物王国。 这种酶的活性与植物萌发有关，人类 哮喘和关节炎。该酶的活性部位含有一种必需的 铁原子。已经提出了两种不同的LO催化机理。 第一个涉及基于自由基的机制，在该机制中，活性中心铁 离子将亚油酸(LA)的1，4-二烯氧化成脂肪酸自由基。 第二种机制是利用铁离子来促进质子的去质子化。 通过协调产生的碳化作用形成基材 有机金属中间体。因此，这两种机制的关键区别在于 是铁中心的作用，以及它是如何激活质子提取的。 这项建议涉及光谱和动力学研究，目的是 两者活性部位铁的结构和功能的研究 大豆和人类脂肪氧合酶。此外，还有一些活动站点 旨在影响铁和它的电子结构的突变体 将对反应性进行研究。光谱技术(例如，EPR，MCD， EXAFS)将用于确定铁配位和/或 电子环境。这些研究将与动力学相关 解决结构变化对 反应性以及pH、粘度和同位素的影响 替换。 获得的信息将被用来确定一个统一的 铁中心在催化氧化中的结构和功能 反应。

DESCRIPTION: Lipoxygenases (LOs) are enzymes that catalyze the incorporation of dioxygen into 1,4-cis,cis-pentadiene containing fatty acids, and are widely distributed throughout the plant and animal kingdoms. The activity of the enzyme has been implicated in plant germination, human asthma and arthritis. The active site of the enzyme contains an essential iron atom. Two distinct mechanisms for LO catalysis have been proposed. The first involves a radical based mechanism in which the active site ferric ion oxidizes the 1,4-diene of linoleic acid (LA) to a fatty acid radical. The second mechanism uses the ferric iron to facilitate de-protonation of the substrate by coordinating the resulting carbonation to form an organometallic intermediate. Thus, the key difference in the two mechanisms is the role of the iron center and how it activates the proton abstraction. This proposal involves spectroscopic and kinetic studies aimed at investigating the structure and function of the active site iron in both soybean and human lipoxygenases. In addition, a number of active site mutants designed to affect the electronic structure of the iron and its reactivity will be studied. Spectroscopic techniques (e.g., EPR, MCD, EXAFS) will be used to determine the changes in the iron coordination and/or electronic environment. These studies will be correlated to kinetic investigations that address the effect of the structural changes on reactivity as well as the effects of pH, viscosity and isotopic substitutions. The information obtained will be used to determine a unified theory of the structure and function of the Fe center in catalyzing the oxidation reactions.