Kucera I, Skládal P
Department of Biochemistry, Faculty of Science, Masaryk University, Brno, Czechoslovakia.
Gen Physiol Biophys. 1990 Oct;9(5):501-17.
Three sites of inhibitory action of hydroxylamine were identified in the respiratory chain of anaerobically grown bacterium Paracoccus denitrificans. Terminal oxidases were blocked at concentrations of 10(-4) to 10(-3) mol.l-1, and the inhibitor competed with artificial donor of electrons N, N, N', N'-tetramethyl-l, 4-phenylenediamine. In the anaerobic part of the respiratory chain inhibition of nitrite reductase and apparently also nitric oxide reductase occurred, resulting in the increased accumulation of nitric oxide during denitrification. These effects together with the inhibition of terminal oxidases by nitric oxide are probably realized through switching the electron flow from oxygen to nitrogen terminal acceptors in the presence of hydroxylamine. By means of difference spectroscopy, the respiratory inhibitor mucidin and a cytochrome c-deficient mutant of Paracoccus denitrificans, hydroxylamine could be shown to serve also as a terminal acceptor of the cytochrome c region. Reduction of hydroxylamine to ammonia was at the same time accompanied by the formation of transmembrane electrical gradient. Hydroxylamine reductase was purified 123-fold from the periplasmatic cell fraction by FPLC; the product obtained showed the features of respiratory nitrite reductase of the cytochrome cd1 type.
在厌氧生长的反硝化副球菌的呼吸链中确定了羟胺的三个抑制作用位点。在浓度为10^(-4)至10^(-3) mol·l^(-1)时，末端氧化酶被阻断，并且该抑制剂与电子人工供体N，N，N'，N'-四甲基-1,4-苯二胺竞争。在呼吸链的厌氧部分，亚硝酸还原酶以及显然还有一氧化氮还原酶受到抑制，导致反硝化过程中一氧化氮积累增加。这些效应以及一氧化氮对末端氧化酶的抑制作用可能是通过在存在羟胺的情况下将电子流从氧末端受体切换到氮末端受体来实现的。通过差示光谱法、呼吸抑制剂粘菌素和反硝化副球菌的细胞色素c缺陷型突变体，表明羟胺也可作为细胞色素c区域的末端受体。羟胺还原为氨的同时伴随着跨膜电位梯度的形成。通过快速蛋白质液相色谱法从周质细胞部分将羟胺还原酶纯化了123倍；所得产物显示出细胞色素cd1型呼吸亚硝酸还原酶的特征。
