Untersuchungen zum Reaktionsmechanismus einer pflanzlichen Oxidase, von Monooxygenasen und von Modellverbindungen für Dioxygenasen

植物氧化酶、单加氧酶和双加氧酶模型化合物的反应机制研究

• 批准号: 5177542
• 负责人: Professor Dr. Bernt Krebs
• 金额: --
• 依托单位: Institut für Anorganische und Analytische Chemie
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 1999
• 资助国家: 德国
• 起止时间: 1998-12-31 至 2003-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/5177542?language=en
• 关键词: Untersuchungen; zum; Reaktionsmechanismus; einer; pflanzlichen

The main objects of research in this proposal are the three metalloenzymes catechol oxidase, tyrosinase, and catechol dioxygenase. The aim of this project is to investigate the possible existence of radical reaction pathways for each of these three enzymes. Catechol oxidase and tyrosinase are related dinuclear copper enzymes which catalyze the oxidation of monophenols or o-diphenols to o-quinones. Our group has determined the X-ray structure of catechol oxidase from sweet potato. The structure which involves an unusual covalent cysteine-histidine bond close to the active site indicates the possibility of a radical reaction pathway. This hypothesis shall be investigated by EPR measurements of the apoenzyme as well as by other spectroscopic methods and kinetic experiments. Different tyrosinases will also be isolated and crystallized. A full single crystal X-ray structure determination is also intended in order to compare these two dicopper enzymes. The proposed mechanism for the reaction pathway of tyrosinase involving radical intermediates will be investigated in detail. Catechol dioxygenase is a non-heme iron enzyme catalyzing the oxidative cleavage of the aromatic rings of catechols. The hypothetical mechanism of this enzyme involves short-lived alkyl-peroxy radicals. From fast kinetic studies on the enzyme and on biomimetic model complexes intermediate semiquinone species shall be trapped by radicals such as NO in order to get insight into the mechanism.

这项建议的主要研究对象是邻苯二酚氧化酶、酪氨酸酶和儿茶酚双加氧酶这三种金属酶。这个项目的目的是调查这三种酶中每一种都可能存在的自由基反应途径。邻苯二酚氧化酶和酪氨酸酶是催化单酚或邻苯二酚氧化为邻苯二酚的双核铜酶。本课题组已经确定了红薯中儿茶酚氧化酶的X射线结构。活性部位附近含有一个不寻常的共价半胱氨酸-组氨酸键的结构表明了自由基反应途径的可能性。这一假说应通过脱辅酶的EPR测量以及其他光谱方法和动力学实验进行验证。不同的酪氨酸酶也将被分离和结晶。为了比较这两种双铜酶，还打算进行全单晶X-射线结构测定。酪氨酸酶涉及自由基中间体的反应途径的提出的机理将被详细研究。儿茶酚双加氧酶是一种催化邻苯二酚芳香环氧化裂解的非血红素铁酶。这种酶的假想机制涉及短命的烷基过氧基。通过对酶的快速动力学研究和仿生模型络合物的研究，中间体半喹酮物种将被NO等自由基捕获，以深入了解其机理。