A Voltammetric Study of Nitrogenase Catalysis Using Electron Transfer Mediators

Nitrogenase catalyzes the reduction of an array of small molecules, including N2 to NH3, by delivering electrons and protons to substrates bound to the active site metal cluster FeMo-cofactor. A challenge in describing the mechanism of nitrogenase-catalyzed reduction reactions is quantifying electron flow through the enzyme to different substrates. In this study, a mediated cyclic voltammetry approach was developed that provides a quantitative analysis of electron flow through nitrogenase. Conditions were optimized to reveal the catalytic reaction rate-limiting step. Analysis of the current response by an electrochemical approach yielded a catalytic rate constant (kcat) of 14 s–1, consistent with earlier studies. The current approach was used to resolve a long-standing conundrum in nitrogenase research, the apparent inhibition of electron flow through nitrogenase with increasing partial pressures of N2. It was demonstrated using this voltammetric approach in the absence of the reductant dithionite that to...