Abstract Enzymatic electrosynthesis has gained more and more interest as an emerging green synthesis platform, particularly for the fixation of CO2. However, the simultaneous utilization of CO2 and a nitrogenous molecule for the enzymatic electrosynthesis of value‐added products has never been reported. In this study, we constructed an in vitro multienzymatic cascade based on the reductive glycine pathway and demonstrated an enzymatic electrocatalytic system that allowed the simultaneous conversion of CO2 and NH3 as the sole carbon and nitrogen sources to synthesize glycine. Through effective coupling and the optimization of electrochemical cofactor regeneration and the multienzymatic cascade reaction, 0.81 mM glycine was yielded with a highest reaction rate of 8.69 mg L−1 h−1 and faradaic efficiency of 96.8 . These results imply a promising alternative for enzymatic CO2 electroreduction and expand its products to nitrogenous chemicals.
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