The chemical evolution of a nitrogenase model, XXIV. Correlational analysis of effects of organic acids on in vitro MoFe-protein substrate reduction activities

Among 29 organic acids whose effects on the in vitro enzymatic substrate reducing activity on the MoFe-protein of nitrogenase have been investigated, 12 acids stimulate the in vitro C2H2-reducing activities in proportion to the H+-reducing activities. A second group comprised of eight acids significantly stimulates the H+-reducing activity but has only modest stimulatory effects on C2H2-and N2-reducing activities. A third group of nine acids causes only slight increases of MoFe-protein substrate reducing activities. The stimulatory effects of acids on MoFe-protein substrate reducing activity depend on their mode of interaction with molybdenum. Hydroxycarboxylic acids acting as bidentate ligands such as homocitric acid and its derivatives leave a sufficient number of molybdenum coordination sites available for interactions with the substrates, they have the highest stimulatory effects both on the C2H2-and N2-reducing activities, and their H+-reducing activities are not inhibited by CO. Acids acting as tridentate ligands, which include citric acid, have weaker stimulatory effects on the C2H2-and especially on the N2-reducing activities, and CO inhibits their H+-reducing activity. Whereas with the first group of acids the C2H2-reducing activities are linearly correlated with H+-reducing activities, the N2-reducing activities are directly correlated with H+-reducing activities in the presence of CO, and the association is exponential rather than linear. This exponential dependence is consistent with a stepwise mechanism of nitrogen reduction via diazene and hydrazine as the intermediates, the latter blocking one molybdenum coordination site prior to its reduction to NH3. In the reduction of C2H2 to C2H4, no such blockage occurs as product C2H4 does not accumulate at the active site and is not reduced further.