Mossbauer Studies of Nitrite Reductase and Vanadium-containing Nitrogenase

生物物质是由能量和化学元素的大规模循环维持的，而生物材料中最基本的元素之一是氮。因此，研究控制生物圈中氮状况的许多不同的生物过程是至关重要的。固氮酶和亚硝酸盐还原酶是氮循环中催化关键步骤的两种酶。固氮酶催化大气中的氮素转化为氨(固氮)，许多微生物利用固氮酶将氮吸收到生命系统中。硝酸还原酶将亚硝酸盐还原为氨和各种硝酸盐还原细菌，以及一些硫酸盐还原细菌，用于终端呼吸，并将氮异化回物理世界。因此，对这些酶的结构和功能的研究具有基本的重要性，并应提供与了解我们的生活环境有关的信息。在这项应用中，Huynh博士建议对以下酶进行穆斯堡尔研究：1.棕色固氮杆菌固氮酶系统的钒-铁蛋白。2.脱硫弧菌的亚硝酸还原酶。钒-铁蛋白和亚硝酸盐还原酶都是含有多个铁中心的复杂酶，穆斯堡尔谱可以最有效地对其进行研究。建议的实验包括(I)天然酶的特性，(Ii)氧化还原滴定研究，(Iii)pH依赖性研究和(Iv)底物和配体结合效应的检测。穆斯堡尔的结果将与从其他物理生化方法获得的数据相关联，包括光学和电子顺磁共振技术。Huynh博士希望全面描述这些酶中的铁中心，获得物理和生化性质，揭示这些铁中心在其酶功能中所起的作用，并加深我们对生物固氮和亚硝酸盐还原机制的理解。

Living matters are sustained by large-scale cycles of energy and chemical elements, and one of the most essential elements in biological materials is nitrogen. To study the many different biological processes which govern the nitrogen status in the biosphere is, therefore, of vital importance. Nitrogenase and nitrite reductase are two enzymes which catalyze key steps in the nitrogen cycle. Nitrogenase catalyzes the conversion of atmospheric dinitrogen into ammonia (nitrogen fixation) and many microorganisms use it to assimilate nitrogen into the living system. Nitrate reductase reduces nitrite to ammonia and a variety of nitrate-reducing bacteria, as well as some sulfate reducing bacteria, use it for terminal respiration and dissimilates nitrogen back to the physical world. Structural and functional studies of these enzymes are therefore of fundamental importance and should provide information pertinent to the understanding of our living environment. In this application, Dr. Huynh proposes to perform Mossbauaer studies of the following enzymes: 1. the vanadium-iron protein of the nitrogenase enzyme system from Azotobacter vinelandii. 2. the nitrite reductase from Desulfovibrio desulfuricans. Both the vanadium-iron protein and nitrite reductase are complex enzymes containing multiple iron centers which can be most effectively studied by Mossbauer spectroscopy. The proposed experiments include (i)characterizations of the native enzymes, (ii)redoxtitration studies, (iii) pH- dependence investigations and (iv)examinations of substrate- and ligand-binding effects. The Mossbauer results will be correlated with data obtained form other physico-biochemical methods, including optical and electron paramagnetic resonance techniques. Dr. Huynh hopes to fully characterize the iron centers in these enzymes, to obtain physical and biochemical properties, to reveal the roles these iron centers play in their enzymatic functions and to enhance our understanding of the mechanisms of biological nitrogen fixation and nitrite reduction.