Guth J H, Burris R H
Biochemistry. 1983 Oct 25;22(22):5111-22. doi: 10.1021/bi00291a010.
We have investigated the inhibition by H2 (D2) of NH3 formation by nitrogenase from Klebsiella pneumoniae and have confirmed that the inhibition is competitive vs. N2. D2 inhibits NH3 formation by diverting nitrogenase from production of NH3 to production of HD (one electron per HD). By careful exclusion of N2 from the reaction mixture, we have been able to place an upper limit on N2-independent HD formation by nitrogenase, under 1 atm of D2, at 1% of the total electron flux. Formation of NH3 and formation of HD were inhibited identically by CO. We observed that as the ratio of dinitrogenase to dinitrogenase reductase is increased, the ratio of HD formed to NH3 formed rises, and D2 becomes a stronger inhibitor of N2 reduction. This may be caused in part by an accompanying increase that is observed in the Km of nitrogenase for N2. We propose a model for D2 inhibition of NH3 formation in which D2 and N2 complete for the same form of nitrogenase. According to our proposal, when N2 reacts with nitrogenase, either N2 reduction proceeds to completion if H2 (D2) is absent or, if D2 already is bound to nitrogenase, N2 reduction is aborted and two molecules of HD are produced at the net expense of one electron per HD. Key consequences of the model are that it predicts that H2 (D2) is a competitive inhibitor of NH3 formation and that the apparent Km (N2) for formation of HD and NH3 may differ.
我们研究了H2（D2）对肺炎克雷伯菌固氮酶形成NH3的抑制作用，并证实这种抑制作用相对于N2是竞争性的。D2通过将固氮酶从NH3的产生转向HD的产生（每个HD一个电子）来抑制NH3的形成。通过小心地从反应混合物中排除N2，我们得以确定在1个大气压的D2下，固氮酶在不依赖N2的情况下形成HD的上限，其占总电子通量的1%。CO对NH3的形成和HD的形成具有相同程度的抑制作用。我们观察到，随着固氮酶与固氮酶还原酶的比例增加，形成的HD与形成的NH3的比例上升，并且D2成为N2还原的更强抑制剂。这可能部分是由于观察到固氮酶对N2的Km值随之增加所致。我们提出了一个D2抑制NH3形成的模型，其中D2和N2竞争相同形式的固氮酶。根据我们的提议，当N2与固氮酶反应时，如果不存在H2（D2），N2还原会进行到底；或者，如果D2已经与固氮酶结合，N2还原就会中止，并且以每个HD净消耗一个电子为代价产生两个HD分子。该模型的关键结果是，它预测H2（D2）是NH3形成的竞争性抑制剂，并且形成HD和NH3的表观Km（N2）可能不同。
