Extracellular Electron Transfer Enables Cellular Control of Cu(I)-Catalyzed Alkyne–Azide Cycloaddition

[Image: see text] Extracellular electron transfer (EET) is an anaerobic respiration process that couples carbon oxidation to the reduction of metal species. In the presence of a suitable metal catalyst, EET allows for cellular metabolism to control a variety of synthetic transformations. Here, we report the use of EET from the electroactive bacterium Shewanella oneidensis for metabolic and genetic control over Cu(I)-catalyzed alkyne–azide cycloaddition (CuAAC). CuAAC conversion under anaerobic and aerobic conditions was dependent on live, actively respiring S. oneidensis cells. The reaction progress and kinetics were manipulated by tailoring the central carbon metabolism. Similarly, EET-CuAAC activity was dependent on specific EET pathways that could be regulated via inducible expression of EET-relevant proteins: MtrC, MtrA, and CymA. EET-driven CuAAC exhibited modularity and robustness in the ligand and substrate scope. Furthermore, the living nature of this system could be exploited to perform multiple reaction cycles without regeneration, something inaccessible to traditional chemical reductants. Finally, S. oneidensis enabled bioorthogonal CuAAC membrane labeling on live mammalian cells without affecting cell viability, suggesting that S. oneidensis can act as a dynamically tunable biocatalyst in complex environments. In summary, our results demonstrate how EET can expand the reaction scope available to living systems by enabling cellular control of CuAAC.