Inhibition of nitrogenase-catalyzed ammonia formation by hydrogen

Abstract

Investigation of the inhibition by H2 (D2) of NH3 formation by nitrogenase from Klebsiella pneumoniae confirmed that the inhibition is competitive vs. N2. D2 inhibits NH3 formation by diverting nitrogenase from production of NH3 to production of HD (1 electron/HD). By careful exclusion of N2 from the reaction mixture, it was possible 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 are inhibited identically by CO. 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. A model is proposed for D2 inhibition of NH3 formation in which D2 and N2 complete for the same form of nitrogenase. According to this 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 2 molecules of HD are produced at the net expense of 1 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.