The mechanism of nitrogenase. Computed details of the site and geometry of binding of alkyne and alkene substrates and intermediates.

The chemical mechanism by which the enzyme nitrogenase effects the remarkable reduction of N(2) to NH(3) under ambient conditions continues to be enigmatic, because no intermediate has been observed directly. Recent experimental investigation of the enzymatic consequences of the valine --> alanine modification of residue alpha-70 of the component MoFe protein on the reduction of alkynes, together with EPR and ENDOR spectroscopic characterization of a trappable intermediate in the reduction of propargyl alcohol or propargyl amine (HCC[triple bond]C-CH(2)OH/NH(2)), has localized the site of binding and reduction of these substrates on the FeMo-cofactor and led to proposed eta(2)-Fe coordination geometry. Here these experimental data are modeled using density functional calculations of the allyl alcohol/amine intermediates and the propargyl alcohol/amine reactants coordinated to the FeMo-cofactor, together with force-field calculations of the interactions of these models with the surrounding MoFe protein. The results support and elaborate the earlier proposals, with the most probable binding site and geometry being eta(2)-coordination at Fe6 of the FeMo-cofactor (crystal structure in the Protein Database), in a position that is intermediate between the exo and endo coordination extremes at Fe6. The models described account for (1) the steric influence of the alpha-70 residue, (2) the crucial hydrogen bonding with Nepsilon of alpha-195(His), (3) the spectroscopic symmetry of the allyl-alcohol intermediate, and (4) the preferential stabilization of the allyl alcohol/amine relative to propargyl alcohol/amine. Alternative binding sites and geometries for ethyne and ethene, relevant to the wild-type protein, are described. This model defines the location and scene for detailed investigation of the mechanism of nitrogenase.