Refinement of the Electrochemical Description of the Nitrogenase Cofactor: Ground States, Activation and N2 Selectivity from Quantum Monte Carlo

Biological nitrogen (N2) fixation by the metalloenzyme nitrogenase is the main enzymatic route which generates ammonia under ambient conditions. Yet, quantum mechanical description of this process remains challenging due to inaccurate energetics of number of close lying states and reaction pathways. Here, we benchmark Density Functional Theory (DFT) with Hubbard U correction against explicitly correlated many‑body quantum Monte Carlo calculations for key states of the iron molybdenum-cofactor (FeMo-co). Using this combined approach, we find the broken-symmetry (BS) 7 type magnetic ordering as the most stable native state (E0) and a triply protonated state (E3) as the electrochemical resting state of the cofactor. We further show that 2 activation of the cofactor proceeds through H2S/HS- unsealing, with an onset potential of -0.88 V vs RHE, close to the experimental ATP/ADP potential (-0.7 V vs RHE) at pH 7. DFT+U revised thermodynamics of the N2 reduction reaction (N2RR) indicate *N2 protonation and S-vacancy unsealing to be the main bottlenecks. Combined energetics of these steps place nitrogenase close to the top of the N2RR volcano within ~0.2 eV of the ATP/ADP experimental onset. Finally, we also extract the adiabatic proton transfer barriers, which suggest that the hydrogen evolution reaction is kinetically hindered relative to N2RR due to the greater distance of the water proton donors from the Fe-Fe active site. Ultimately, this work underscores the importance of the correlated treatment of Fe metal centers and their propensity to unseal and activate with the evolution of hydrogen sulfide ions as the key to the cofactor’s activity towards N2RR.