Cofactor-Dependent Amine Oxidations

说明(申请人提供)：含铜的胺氧化酶(CAO)包括所有生物中普遍存在的一类酶。在植物和哺乳动物中，CAO在代谢生物胺方面发挥着重要作用，这些生物胺参与了生长、细胞分裂、分化和应激反应，在某些情况下，代谢具有细胞毒性后果。所有CAO都利用翻译后从活性部位酪氨酸残基衍生的苯醌辅因子来调节伯胺到醛的转氨基转换。在过去的资助期间，使用辅因子模型的研究启发了新类型的酶激活抑制机制的发现。这些研究，以及其他传统失活策略的发展，已经导致了非常有希望的抑制剂选择性。所实现的选择性不仅区别于黄素和苯二酚依赖的酶，而且区别于具有不同底物结构偏好的一系列CAO。自上一次更新以来，人类CAO的两大类的遗传学基础已经被揭示：(1)依赖三羟基苯丙氨酸二酚(TPQ)的类AOC 1(人肾二胺氧化酶)、AOC 2和AOC 3，后者与人血管黏附蛋白-1(HVAP-1)重合；(2)除了经典的LOX外，还有4种赖氨酰氧化酶样蛋白(LOXL)。在下一个赠款期间，将更多地关注精制人类酶的抑制剂选择性，特别是肾脏/胎盘二胺氧化酶、可溶性和组织结合的氨基脲敏感的单胺氧化酶和赖氨酰氧化酶。新的目标包括开发检测和定位哺乳动物CAO的灵敏荧光分析方法，探索新的抑制剂选择性和前药结构，利用X射线结晶学信息(通过合作)改进通过分子模拟构建的底物和抑制剂结合模型，以及使用光谱和质谱学来确定不可逆抑制剂作用的结构基础。此外，还将完成对(TPQ)辅因子生物发生性质的研究。 这项研究与公共卫生的相关性 随着人们对铜胺氧化酶在炎症、免疫调节、关节炎和动脉粥样硬化的纤维化以及糖尿病和心血管疾病的晚期并发症中的关键生理作用的认识，铜胺氧化酶已成为重要的药物靶标。因此，重要的是要从机理和底物识别的结构基础以及可在最终药物开发中起先导作用的抑制剂的作用方面了解这些酶的功能。

DESCRIPTION (provided by applicant): The copper-containing amine oxidases (CAOs) comprise a ubiquitous class of enzymes in all living organisms. In plants and mammals, CAOs play important roles in metabolizing biogenic amines involved in growth, cell division, differentiation, and the stress response, where metabolism in some cases has cytotoxic consequences. All CAOs utilize a quinone cofactor derived postranslationally from an active-site tyrosine residue to mediate a transaminative conversion of primary amines to aldehydes. Studies using cofactor models in the past grant periods have inspired the discovery of new types of enzyme-activated mechanisms of inhibition. These studies, along with development of other traditional inactivation strategies, have resulted in highly promising degrees of inhibitor selectivity. The selectivity being achieved is discriminating not only between the flavin- and quinone-dependent enzymes, but also among a range of CAOs that have different substrate-structure preferences. Since the time of the last renewal, the genetic basis of the two general classes of human CAOs has been revealed: (i) the trihydroxyphenylalanine quinone (TPQ)-dependent class AOC 1 (the human kidney diamine oxidase), AOC 2, and AOC 3, the latter coincidental with the human vascular adhesion protein-1 (HVAP-1), and (ii) four lysyl oxidase-like (LOXL) proteins in addition to classical LOX. In the next grant period, there will be increased focus on refining inhibitor selectivity for the human enzymes, particularly the kidney/placental diamine oxidase, the soluble and tissue-bound semicarbazide- sensitive monoamine oxidases, and lysyl oxidase. The new aims include the development of sensitive fluorometric assays for detecting and localizing mammalian CAOs, an exploration of new inhibitor selectivity and prodrug constructs, the use of x-ray crystallographic information (through collaboration) for refinement of substrate and inhibitor binding models constructed by molecular modeling, and the use of spectroscopy and mass spectrometry to identify the structural basis of irreversible inhibitor action. Studies on the nature of biogenesis of the (TPQ) cofactor will additionally be completed. RELEVANCE OF THIS RESEARCH TO PUBLIC HEALTH The copper amine oxidases have become important pharmaceutical targets on account of the emerging recognition of their key physiological roles in aspects of inflammation, immune modulation, fibrosis in arthritis and atherosclerosis, and in late-stage complications of diabetes and cardiovascular disease. It is thus important to understand the functioning of these enzymes, both from a mechanistic view and in terms of the structural basis of substrate recognition and the action of inhibitors that can serve a leads in eventual drug development.