Structural Studies of Alternating-site Reactivity in Nitrogenase-like Oxidoreductases

项目摘要/摘要 自然界利用蛋白质变构来控制化学反应中的远程电子流动 终生如此。金属和金属团簇往往是促进这种电子转移的关键成分。一个 已知的用于在金属酶中门控电子流的变构方法是交替位置反应性， 其中，当构象时，反应性从酶的一半切换到另一半 改变。氧化还原酶具有核心的α-2-β-2异四聚体结构，是铁-硫簇 利用交替位置反应性的金属酶，可以作为一个无价的模型 定义金属酶变构所需结构基序和机制的系统 监管。符合NIH NIGMS药理学、生理学和生化化学 “提高对基本生物过程和分子水平的理解”的使命 发现它们的控制方法“，我们建议研究两种这样的氧化还原酶，即 固氮酶样暗作用叶绿素氧化还原酶(DPOR)和叶绿素 氧化还原酶(COR)，以确定远程相关运动对交替运动的贡献。 使用结构生物学工具的现场反应性。我们将使用电子显微镜和小角 不对称酶构象中间体的X射线散射检测技术 构象。

Project Summary / Abstract Nature uses protein allostery to control long-range electron flow in chemical reactions essential for life. Metals and metal-clusters are often key components facilitating this electron transfer. A known allosteric method for gating electron flow in metalloenzymes is alternating-site reactivity, wherein reactivity switches from one half of the enzyme to the other upon conformational changes. Oxidoreductases, with a core α2β2 heterotetrameric structure, are iron-sulfur cluster metalloenzymes that employ alternating-site reactivity and can serve as an invaluable model system to define the structural motifs and mechanisms required for metalloenzyme allosteric regulation. In line with the NIH NIGMS Pharmacology, Physiology, and Biochemical Chemistry mission of “improving molecular level understanding of fundamental biological processes and discovering approaches to their control”, we propose to study two such oxidoreductases, the nitrogenase-like dark-operative chlorophyllide oxidoreductase (DPOR) and chlorophyllide oxidoreductase (COR), to identify how long-ranged correlated motions contribute to alternating- site reactivity using structural biology tools. We will employ electron microscopy and small-angle X-ray scattering techniques to examine enzyme conformational intermediates with asymmetrical conformations.