Mechanistic Studies of Lipoic Acid Synthase

描述(申请人提供)：硫辛酸是必需的 所涉及的几种多酶复合体的含硫辅因子 在初级新陈代谢中。多年来，人们已经知道它是由 直接来自辛酸，但硫插入到 未活化的脂肪酸仍然鲜为人知。最近，一种基因被克隆出来。 它似乎是酶活性的催化核心，即 参与了硫插入反应。推论出的签名主题 该蛋白质的一级序列和初步波谱特征 共同表明这种“硫辛酸合成酶”属于一种新兴的 一类含有铁-硫簇的酶，并且使用 S-腺苷-L-蛋氨酸作为生成碳心自由基的方法 酶反应中的中间体。S-腺苷-L-蛋氨酸和蛋氨酸的作用 这类酶中的铁-硫团簇代表着与 他们传统上在生物化学教科书中被分配的角色。 传统上，铁-硫团簇被认为是电子转移 试剂或Lewis酸催化剂，而S-腺苷-L-蛋氨酸仍 被认为主要是一种细胞甲基化试剂。建议进行的实验 在这里试图揭示与这些相关的“新的”分子酶学 辅因。讨论了几种实验机制，并进行了实验 建议将它们区分开来。 除了硫辛酸作为辅因子参与的酶复合体 能量代谢，已知可以调节糖尿病患者的葡萄糖代谢 II型糖尿病，作为一种普通的细胞抗氧化剂，在许多 其他的事情。有重要证据表明，硫辛酸可以 在哺乳动物细胞中内源性合成。这篇文章中概述的实验 建议将为研究奠定基础，以解决 不能内源性合成硫辛酸会导致 细胞功能。

DESCRIPTION (provided by applicant): Lipoic acid is an essential sulfur-containing cofactor of several multienzyme complexes that are involved in primary metabolism. It has been known for a number of years to be derived directly from octanoic acid, but the mechanism of sulfur insertion into the unactivated fatty acid has remained obscure. Recently, a gene has been cloned which appears to be the catalytic core of the enzymatic activity that is involved in the sulfur insertion reaction. Signature motifs within its deduced primary sequence and initial spectroscopic characterization of the protein collectively indicate that this "lipoic acid synthase" belongs to an emerging class of enzymes that contain iron-sulfur clusters, and that use S-adenosyl-L-methionine as a means of generating carbon-centered radical intermediates in enzymatic reactions. The roles of S-adenosyl-L-methionine and iron-sulfur clusters in this class of enzymes represent departures from the roles that they have traditionally been assigned in biochemical textbooks. Iron-sulfur clusters have traditionally been thought of as electron transfer agents or Lewis acid catalysts, while S-adenosyl-L-methionine is still considered to be mainly a cellular methylating agent. The experiments proposed herein seek to reveal the "new" molecular enzymology associated with these cofactors. Several experimental mechanisms are discussed, and experiments are proposed that will distinguish among them. Aside from the involvement of lipoic acid as a cofactor in enzyme complexes of energy metabolism, it is known to modulate glucose metabolism in patients with type II diabetes and to serve as a general cellular antioxidant, among many other things. There is significant evidence that lipoic acid can be endogenously synthesized in mammalian cells. Experiments outlined in this proposal will lay a foundation upon which studies to address whether the inability to synthesize lipoic acid endogenously can lead to a compromising of cellular function.