Photoinduced Reductive Elimination of H from the Nitrogenase Dihydride (Janus) State Involves a FeMo-cofactor-H Intermediate.

N reduction by nitrogenase involves the accumulation of four reducing equivalents at the active site FeMo-cofactor to form a state with two [Fe-H-Fe] bridging hydrides (denoted E(4H), the Janus intermediate), and we recently demonstrated that the enzyme is activated to cleave the N≡N triple bond by the reductive elimination (re) of H from this state. We are exploring a photochemical approach to obtaining atomic-level details of the re activation process. We have shown that, when E(4H) at cryogenic temperatures is subjected to 450 nm irradiation in an EPR cavity, it cleanly undergoes photoinduced re of H to give a reactive doubly reduced intermediate, denoted E(2H), which corresponds to the intermediate that would form if thermal dissociative re loss of H preceded N binding. Experiments reported here establish that photoinduced re primarily occurs in two steps. Photolysis of E(4H) generates an intermediate state that undergoes subsequent photoinduced conversion to [E(2H) + H]. The experiments, supported by DFT calculations, indicate that the trapped intermediate is an H complex on the ground adiabatic potential energy suface that connects E(4H) with [E(2H)* + H]. We suggest that this complex, denoted E(H; 2H), is a thermally populated intermediate in the catalytically central re of H by E(4H) and that N reacts with this complex to complete the activated conversion of [E(4H) + N] into [E(2N2H) + H].