Enzymology of the Hydroxylamine Oxidizing System of Nitrosomonas

在土壤中，肥料氨被氧化成亚硝酸盐（NH3+1.5 02-HNO 2 +H20）的硝化作用 欧洲亚硝化单胞菌 亚硝酸盐被硝化菌氧化成 硝酸盐是非农业地区食物蛋白质氮的主要来源， 豆科作物 因此，大多数人最初的转变 食物氮是由亚硝化单胞菌进行的。 大 经济上的重要性是土壤中氮的损失，如氮气体 由亚硝化单胞菌在NH3氧化过程中产生，或由 通过微生物减少硝酸盐。 控制 反硝化作用造成的农业损失试图通过使用 亚硝化单胞菌属氨氧化的抑制剂。 这 有助于加深对 氨氧化 NH3被O2氧化成NH 2 OH，然后被氧化 （NH 2 OH +H2）--HNO 2 + 4 e-+4H+）通过酶羟胺 氧化还原酶，“HAO.“HAO是一个大的（200 Kd）， 异常复杂的血红素蛋白（24个C血红素和3个底物 结合“P460”基团）。 两个电子从HAO通过， 在包括四血红素的电子传递链中， 细胞色素c554和其他未知成分， 末端氧化酶 另外两个电子从HAO传递到 氨单加氧酶。 该项目的目标包括： 亚基结构和氨基酸序列的测定 HAO的P460的环结构和HAO的P460的结构 的底物结合位点和鉴定的 HAO电子出口部位;氨基酸的测定 细胞色素c的电子进出位点 554;以及用末端氧化酶重建HAO。 该方法涉及酶学，蛋白质化学，光学 以及EPR光谱快速动力学、克隆和DNA测序。

In soils, fertililzer ammonia is oxidized to nitrite (NH3+1.5 02--HNO2+H20) by the chemautotrophic nitrifying bacterium Nitrosomonas europaea. Nitrite is oxidized by Nitrobacter to nitrate, a primary source of N for food protein in non- leguminous crops. Thus the initial transformation of most food ntirogen is carried out by Nitrosomonas. Of great economic importance is loss of N in soils, as N-gases produced by Nitrosomonas during oxidation of NH3 or by reduction of nitrate by denitrifying bacteria. Control of agricultural loss by denitrification is attempted by the use of inhibitors of ammonia oxidation by Nitrosomonas. This lends significance to improved knowledge of the mechanism of ammonia oxidation. NH3 is oxygenated by O2 to NH2OH which is then oxidized (NH2OH+H2)--HNO2+4e-+4H+) by the enzyme hydroxcylamine oxidoreductase, "HAO." HAO is a large ( 200 Kd) and unusually complex hemoprotein ( 24 c hemes and 3 substrate binding "P460" groups). Two electrons pass from HAO, in an electron transport chain which includes the tetraheme cytochrome c554 and other unknown components, to a membrane terminal oxidase. Two additional electrons pass from HAO to the ammonia monoxygenase. Goals of the project include: determination of the subunit structure and amino acid sequence of HAO, the ring structure of P460 of HAO and the structure of the substrate binding site and identification of the electron exit site of HAO; determination of the amino acid sequence and electron entry and exit sites of cytochrome c- 554; and reconstitution of HAO with the terminal oxidase. The approach involves enzymology, protein chemistry, optical and EPR spectroscopy rapid kinetics, cloning and DNA sequencing.