Mechanisms of Flavoproteins

描述（由申请人提供）：黄蛋白催化广泛的生物关键反应。其中许多都涉及碳氧键或碳氮键的双电子氧化。尽管这些反应很重要，但这些酶的作用机制仍然存在争议。在下一个资助期，我们建议将重点放在氧化碳和氮之间键的黄酶上。我们建议研究两种参与多胺代谢的酶：优先氧化乙酰化精胺和亚精胺的多胺氧化酶和优先氧化未乙酰化精胺的精胺氧化酶。多胺对细胞的生长至关重要，多胺氧化酶和精胺氧化酶的替代底物可以防止癌细胞的生长。本文将研究这两种酶对底物特异性的催化机制和结构基础。我们还将继续研究硝基烷烃氧化酶。这种酶是一种不寻常的黄素蛋白，它通过形成碳离子中间体来氧化硝基烷烃的碳-氮键。该酶可作为黄素蛋白催化质子从碳中去除的模型。研究这三种酶的方法将结合稳态和快速反应动力学，包括动力学同位素效应，以及结构方法。在硝基烷烃氧化酶的情况下，非酶反应也将进行研究，以深入了解酶提高质子提取速率的机制。公共卫生相关性：多胺氧化酶和精胺氧化酶都参与多胺的再循环。细胞的生长需要多胺，而破坏多胺代谢的化合物已被证明可以阻止癌细胞的生长。硝基烷烃氧化酶是研究蛋白质脱碳机理的模型酶。

DESCRIPTION (provided by applicant): Flavoproteins catalyze a wide range of biologically critical reactions. Many of these involve the two electron oxidation of a carbon-oxygen or carbon-nitrogen bond. Despite the importance of such reactions, the mechanisms of these enzymes continue to be controversial. In the next grant period we propose to focus on flavoenzymes which oxidize bonds between carbon and nitrogen. We propose to study two enzymes involved in metabolism of polyamines: polyamine oxidase, which preferentially oxidizes acetylated spermine and spermidine, and spermine oxidase, which prefers unacetylated spermine. Polyamines are critical for the growth of cells, and alternate substrates for polyamine oxidase and spermine oxidase prevent growth of cancer cells. The catalytic mechanisms and structural bases for substrate specificity in these two enzymes will be studied. We will also continue study of nitroalkane oxidase. This enzyme is an unusual flavoprotein which oxidizes the carbon-nitrogen bond of nitroalkanes by forming a carbanion intermediate. The enzyme serves as a model for flavoprotein-catalyzed proton removal from carbon. The approaches to all three enzymes will combine steady-state and rapid-reaction kinetics, including kinetic isotope effects, with structural approaches. In the case of nitroalkane oxidase, the nonenzymatic reaction will also be studied to gain insight into the mechanism by which the enzyme increases the rate of proton abstraction. PUBLIC HEALTH RELEVANCE: Polyamine oxidase and spermine oxidase are both involved in recycling of polyamines. Polyamines are needed for the growth of cells, and compounds which disrupt}polyamine metabolism have been shown to prevent the growth of cancer cells. Nitroalkane oxidase is a model enzyme for studying the mechanisms of proteins which remove a proton from carbon.