Nitrogenase Bioelectrocatalysis: ATP-Independent Ammonia Production Using a Redox Polymer/MoFe Protein System

Nitrogenase is the only biological catalyst that is known to be able to convert nitrogen gas to ammonia. In microorganisms, the MoFe catalytic protein of nitrogenase is reduced by a transient Fe protein binding and separate hydrolysis of ATP. However, the requirement of 16 ATP molecules by the Fe protein for the 8 electron transfer is an energy-intense caveat to the enzymatic synthesis of NH₃ and is challenging from an electrochemical perspective. Thus, we report the redox polymer-based ATP-free mediated electron-transfer system of MoFe nitrogenase using cobaltocene-functionalized poly­(allylamine) (Cc-PAA), which is able to reduce the MoFe nitrogenase directly with a low redox potential of −0.58 V vs SHE. An efficient immobilization of MoFe nitrogenase via Cc-PAA allowed for the bioelectrocatalytic reduction of N₃^–, NO₂^–, and N₂ to NH₃. Bulk bioelectrosynthetic experiments produced 7 ± 2 and 30 ± 5 nmol of NH₃ from NO₂^– and N₃^– reduction for 30 min, respectively. In addition, biosynthetic N₂ reduction to NH₃ was confirmed by ¹⁵N₂ labeling experiments with NMR analysis. This mediated electron-transfer approach of the immobilized nitrogenase using the Cc-PAA redox polymer provides a valuable technological basis for scale-up and industrial uses in the future of bioelectrosynthesis.