Understanding non-reducible N₂ in the mechanism of Mo–nitrogenase

In my proposed mechanism of Mo-nitrogenase there are two roles for separate N₂ molecules. One N₂ diffuses into the reaction zone between Fe2 and Fe6 where a strategic gallery of H atoms can capture N₂ to form the Fe-bound HNNH intermediate which is then progressively hydrogenated through intermediates containing HNNH₂, NH and NH₂ entities and then two NH₃ in sequence. The second N₂ can be parked in an N₂-pocket about 3.2 Å from Fe2 or bind end-on at the <i>exo</i> coordination site of Fe2. This second N₂ is outside the reaction zone, not exposed to H atom donors, and so is 'non-reducible'. Here density functional calculations using a 485+ atom model describe the thermodynamics for non-reducible N₂ moving between the N₂-pocket and the <i>exo</i>-Fe2 position, for the resting state and 19 intermediates in the mechanism. The entropy component is estimated and included. The result is that for all intermediates with ligation by H or NH_<i>x</i> at the <i>endo</i>-Fe2 position the free energy for association of non-reducible N₂ at <i>exo</i>-Fe2 is negative. There remains some uncertainty about the status of <i>exo</i>-Fe2-N₂ during the step in which H₂ exchanges with the incoming reducible N₂, where at least two unbound molecules are present. At Fe2 it is evident that attainment of octahedral coordination stereochemistry dominates the binding thermodynamics for non-reducible N₂. Possibilities for experimental support of these computational conclusions are discussed.