Photoinduced Reductive Elimination of H₂ from the Nitrogenase Dihydride (Janus) State Involves a FeMo-cofactor-H₂ Intermediate

N₂ 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 E₄(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 H₂ from this state. We are exploring a photochemical approach to obtaining atomic-level details of the re activation process. We have shown that, when E₄(4H) at cryogenic temperatures is subjected to 450 nm irradiation in an EPR cavity, it cleanly undergoes photoinduced re of H₂ to give a reactive doubly reduced intermediate, denoted E₄(2H)*, which corresponds to the intermediate that would form if thermal dissociative re loss of H₂ preceded N₂ binding. Experiments reported here establish that photoinduced re primarily occurs in two steps. Photolysis of E₄(4H) generates an intermediate state that undergoes subsequent photoinduced conversion to [E₄(2H)* + H₂]. The experiments, supported by DFT calculations, indicate that the trapped intermediate is an H₂ complex on the ground adiabatic potential energy suface that connects E₄(4H) with [E₄(2H)* + H₂]. We suggest that this complex, denoted E₄(H₂; 2H), is a thermally populated intermediate in the catalytically central re of H₂ by E₄(4H) and that N₂ reacts with this complex to complete the activated conversion of [E₄(4H) + N₂] into [E₄(2N2H) + H₂].