Biophysical Studies of Metalloenzymes

题目：金属酶的生物物理研究该项目由生物科学系统计划理事会分子和细胞生物科学部分子生物物理集群和数学与物理科学理事会化学部门生命过程化学计划共同资助。生物有效氮来源于大气氮还原产生的氨，是植物生长中仅次于水的限制性养分。从大约20亿年前氮酶的出现，直到20世纪50年代广泛使用哈伯-博世法化学生产氨肥料，所有生命都通过利用氮酶的生物固氮来获得氮。目前，利用氮酶的生物固氮提供了世界上大约一半的氨。剩下的一半由Haber-Bosch工业流程生产，该流程约占世界总能耗的2%。本研究的目的是揭示氮酶的作用机制。了解氮酶的功能将有可能使氮酶活性的有效利用为人类带来更大的利益，增加粮食产量和减少氨肥工业生产中的能源消耗。此外，科研人员还积极参与各级科学教育培训工作。该项目的第一个目标是通过使用遗传/生化方法分离催化中间体，结合实验室使用电子-核双共振（ENDOR）和电子自旋回波包络调制（ESEEM）光谱来表征反应中间体，进一步完善研究人员阐明的氮酶反应机制。该项目的第二个目标是通过将氢、氘、（14,15）氮、（57）铁和（95）钼的ENDOR/ESEEM光谱研究与量子化学计算相结合，首次将QM/MM方法引入到氮酶中间体的研究中，从而确定整个催化循环过程中酶中间体的结构和电子性质。该项目的第三个目标是研究仿生钼铁配合物，以促进对这些配合物的生物催化潜力的理解。仿生Mo和Fe配合物的EPR/ENDOR特性对我们的氮酶中间体中底物衍生部分的鉴定产生了强大的限制。此外，这些配合物对配位化学有其自身的基础重要性，包括非经典的M-H(2)加合物，它们表现出新的H(2)旋转动力学，也与氢催化和能量储存有关。

Title: Biophysical Studies of MetalloenzymesThis project is jointly funded by the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences in the Directorate for Biological Sciences Systems Program and the Chemistry of Life Processes Program in the Division of Chemistry in the Directorate of Mathematical and Physical Sciences. Bioavailable nitrogen derived from ammonia produced by the reduction of atmospheric nitrogen is second only to water as the limiting nutrient in plant growth. From the appearance of the enzyme nitrogenase approximately two billion years ago, until the wide-spread use of the Haber-Bosch process in the 1950's for the chemical production of ammonia fertilizers, all life obtained nitrogen from biological nitrogen fixation using the nitrogenase enzyme. Currently, biological nitrogen fixation using nitrogenase supplies approximately one-half of the world's ammonia. The remaining half is produced by the Haber-Bosch industrial process, a process that is responsible for approximately 2% of the world's total energy consumption. The goal of this research is to reveal the mechanism of nitrogenase function. Understanding of nitrogenase function will potentially enable the effective utilization of nitrogenase activity for greater human benefit, increasing food production and reducing energy consumption during the industrial production of ammonia fertilizers. In addition, investigators are active in science education and training at all levels.The first goal of this project is to further refine the nitrogenase reaction mechanism elucidated by the investigators through the use of genetic/biochemical approaches for the isolation of catalytic intermediates in combination with the laboratory's use of electron-nuclear double resonance (ENDOR) and electron spin-echo envelope modulation (ESEEM) spectroscopies to characterize the reaction intermediates. A second goal of the project is to determine the structure and electronic properties of enzymatic intermediates throughout the catalytic cycle, by integrating hydrogen, deuterium, (14,15)nitrogen, (57)Fe and (95)Mo ENDOR/ESEEM spectroscopic studies with quantum chemical calculations that for the first time introduce QM/MM methods to the study of nitrogenase intermediates. The third goal of the project is to study the biomimetic molybdenum and iron complexes in order to advance the understanding of the biocatalytic potential of these complexes. EPR/ENDOR characterizations of biomimetic Mo and Fe complexes yield powerful constraints on the identification of substrate-derived moieties in our nitrogenase intermediates. In addition, these complexes have their own, fundamental importance to coordination chemistry that include non-classical M-H(2) adducts that show novel H(2) rotational dynamics and also are relevant for hydrogen catalysis and energy storage.