Thermodynamically Favourable States in the Reaction of Nitrogenase without Dissociation of any Sulfide Ligand

We have used combined quantum mechanical and molecular mechanical (QM/MM) calculations to study the reaction mechanism of nitrogenase, assuming that none of the sulfide ligands dissociates. To avoid the problem that there is no consensus regarding the structure and protonation of the E(4) state, we start from a state where N(2) is bound to the cluster and is protonated to N(2)H(2), after dissociation of H(2). We show that the reaction follows an alternating mechanism with HNNH (possibly protonated to HNNH(2)) and H(2)NNH(2) as intermediates and the two NH(3) products dissociate at the E(7) and E(8) levels. For all intermediates, coordination to Fe6 is preferred, but for the E(4) and E(8) intermediates, binding to Fe2 is competitive. For the E(4), E(5) and E(7) intermediates we find that the substrate may abstract a proton from the hydroxy group of the homocitrate ligand of the FeMo cluster, thereby forming HNNH(2), H(2)NNH(2) and NH(3) intermediates. This may explain why homocitrate is a mandatory component of nitrogenase. All steps in the suggested reaction mechanism are thermodynamically favourable compared to protonation of the nearby His‐195 group and in all cases, protonation of the NE2 atom of the latter group is preferred.