A MECHANISTIC STUDY OF SOME O2 OXIDOREDUCTASES

这项研究的长期目标是详细确定不同的 O2氧化还原酶催化O2的反应。因为这些酶是 许多重要代谢途径所必需的(包括：肌醇 分解代谢、芳香族氨基酸代谢、儿茶酚胺的生物合成和 降解、类固醇激素生物合成、药物代谢等)，它是 我觉得更详细地了解它们的机制可能会导致 新陈代谢控制或减轻许多新陈代谢缺陷三种酶 下面列出的将在即将到来的授权期内进行调查，但 主要重点将放在第一个方面。 猪肾肌醇加氧酶。这种酶负责 在动物体内肌醇分解代谢的第一步，已知的是 缺乏患糖尿病的动物。因为肌醇衍生品现在 已知充当各种激素的第二信使，一项详细的研究 肌醇酶部分负责细胞肌醇状态的酶 这似乎是有根据的。拟议工作的一个主要重点将是确定 并对参与催化的酶基团进行表征，有望获得 关于该酶的详细结构信息。为此，我们计划： 进一步鉴定和开发酶活性的亲和标记 通过以下方法获得该酶的完整氨基酸序列 该酶基因的克隆和测序，鉴定了金属离子和 它的结合部位，有望获得适合于完整X射线的晶体 结构确定等。 猪肝中的4-羟基苯丙酮酸双加氧酶。这种酶，它是 参与所有动物的酪氨酸代谢，参与 几种代谢性疾病。大多数拟议的实验都是为了 鉴定反应中可能的中间体或氨基酸 活性部位的残留物。 牛肾中的D-天冬氨酸氧化酶。这种过氧化物酶被认为是 在控制动物新陈代谢方面起作用。我们的目标是 建议的研究是通过动力学方法确定(特别是使用 停流技术)对这种异常激活的解释 一些效应器产生的单体黄素酶。

The long term goal of this research is to determine in detail how various O2 oxidoreductases catalyze reactions of O2. Since these enzymes are necessary for so many important metabolic pathways (including: inositol catabolism, aromatic amino acid metabolism, catecholamine biosynthesis and degradation, steroid hormone biosynthesis, drug metabolism, etc.), it is felt that a more detailed understanding of their mechanisms could lead to the control or alleviation of many metabolic defects. The three enzymes listed below will be investigated during the coming grant period, but the major emphasis will be on the first. myo-Inositol oxygenase from pig kidney. This enzyme is responsible for the first committed step in inositol catabolism in animals and is known to be deficient in animals with diabetes. Since inositol derivatives are now known to act as second messengers for various hormones, a detailed study of the enzyme that is partially responsible for the inositol status of cells seems warranted. A major focus of the proposed work will be to identify and characterize enzymic groups involved in catalysis, and hopefully obtain detailed structural information about the enzyme. To this end we plan to: further characterize and develop affinity labels for the enzyme's active site residues, obtain the complete amino acid sequence of the enzyme by cloning and sequencing the enzyme's cDNA, characterize the metal ion and its binding site, hopefully obtain crystals suitable for a complete X-ray structure determination, etc. 4-Hydroxyphenylpyruvate dioxygenase from pig liver. This enzyme, which participates in the metabolism of tyrosine in all animals, is involved in several metabolic diseases. Most of the proposed experiments are aimed at identifying, either possible intermediates in the reaction, or amino acid residues at the active site. D-Aspartate oxidase from beef kidney. This peroxisomal enzyme is believed to function in the control of metabolism in animals. Our goal in the proposed research is to determine by kinetic methods (especially employing stopped flow techniques) the explanation for an unusual activation of this monomeric flavoenzyme by some effectors.