Electroenzymatic N2 reduction to ammonia by nitrogenase from Azotobacter vinelandii immobilized on low density graphite electrodes under direct electron transfer regime

Nitrogenases accomplish the energetically challenging reduction of nitrogen to produce ammonia under mild conditions. Co-immobilization of both components of the Mo-nitrogenase from Azotobacter vinelandii on porous low-density graphite electrodes has allowed studying the electroenzymatic ammonia production in absence of any redox mediator. The entrapment of both nitrogenase proteins in an electrode with adequate pore size distribution has allowed direct electron transfer at the electrode while preserving enough mobility for their association and dissociation, which is required for the ATP-dependent catalytic turnover of nitrogen reduction to ammonia. Both cyclic voltammetry and chronoamperometry measurements showed that the electrocatalytic effect of N 2 reduction required the addition of ATP to the electrolyte. Quantification of the ammonia produced was performed by a spectrophotometric method based on the activity of glutamate dehydrogenase. An average value of 15 ± 6 nmol of NH 4 + was determined after 120 min at an applied potential of -0.9 vs. Ag/AgCl (3 M Cl − ) with a faradaic yield of 65 ± 5 %.