The Pathway of Denitrification

反硝化作用是氮循环的关键过程， 控制着植物生长所需的氮量。 它 据估计，高达30%的肥料氮是通过 脱氮作用 硝酸盐被还原成 气态氮的详细情况还不清楚，但亚硝酸盐和 一氧化二氮已被确定为游离中间体， 从NO2 -到N2 O似乎有两条途径，一条是直接的， 另一个通过不。 拟议研究的目的是阐明 NO2 -转化为N2 O的途径。 主要 需要回答的问题是NO2 -如何被血红素cd 1还原 和含铜的亚硝酸盐还原酶，NO是如何被还原成 N2 O的一氧化氮还原酶，什么因素控制， 通过两条途径从NO2 -到N2 O在体内的相对通量。 一 多方面的方法来解决这个问题，结合 化学和生物学研究，并侧重于机制 N2 O的N-N键由此形成。 具体技术， 包括同位素标记实验;动力学 研究;冷冻酶学和光谱学研究的 纯化的酶;和电子显微镜。具体制度， 包括全细胞和无细胞提取物 选择的野生型细菌和合适的突变体， 以及纯化的亚硝酸还原酶和粗一氧化氮 还原酶制剂。

Denitrification is a key process in the nitrogen cycle, which controls the amount of nitrogen available for plant growth. It is estimated that up to 30% of fertilizer nitrogen is lost via denitrification. The pathway by which nitrate is reduced to gaseous nitrogen is still not known in detail, but nitrite and nitrous oxide have been identified as free intermediates, and there appear to be two routes from NO2 - to N2O, one direct and the other via NO. The object of the proposed research is to elucidate the pathway(s) by which NO2 - is converted to N2O. The major questions to be answered are how NO2 - is reduced by the heme cd1 and copper-containing nitrite reductases, how NO is reduced to N2O by nitric oxide reductase, and what factors control the relative flux via the two routes from NO2 - to N2O in vivo. A multifaceted approach to the problem is described, combining chemical and biological studies, and focussing on the mechanism by which the N-N bond of N2O is formed. Specific techniques to be employed include isotopic labelling experiments; kinetics studies; cryoenzymological and spectroscopic studies of the purified enzymes; and electron microscopy. Specific systems to be examined include whole cells and cell-free extracts of selected wild-type denitrifying bacteria and appropriate mutants, as well as purified nitrite reductase and crude nitric oxide reductase preparations.