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

Abstract

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.

Rewiring native post-transcriptional carbon regulators to build multi-layered genetic circuits and optimize engineered microbes for bioproduction

“…We also sought to re-establish similar regulatory capabilities by leveraging the conserved CsrA homologs. We first selected Shewanella oneidensis MR-1 as the bacteria possess unique bioprocessing capabilities of industrial relevance (Partipilo et al 2022 ACS Cent. Sci.…”

Nanotechnology connecting copper metabolism and tumor therapy

“…Therefore, Cu is the catalytic center of a variety of metabolic enzymes, including monoamine oxidase, superoxide dismutase, and ceruloplasmin 7–16 . Cu has a significant role in electron transfer, protein synthesis, purine metabolism, lipid and neuron development, as well as the activation of hemoglobin synthesis and the promotion of iron uptake and use 17–33 …”

“…[7][8][9][10][11][12][13][14][15][16] Cu has a significant role in electron transfer, protein synthesis, purine metabolism, lipid and neuron development, as well as the activation of hemoglobin synthesis and the promotion of iron uptake and use. [17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33] According to several studies, different malignant tumors have higher Cu contents than normal tissues. 34 Cu can stimulate angiogenesis, 35 which is necessary for the growth and metastasis of tumors, by activating angiogenic proteins like angiopoietin (Ang), vascular endothelial growth factor (VEGF), [36][37][38][39] fibroblast growth factor 1 (FGF1), 40 and interleukin-1 (IL-1), and promote angiogenesis.…”

Single-Cell Phenotyping of Extracellular Electron Transfer via Microdroplet Encapsulation

“…We characterized two bacterial species not previously shown to exhibit electrogenic behavior. Our results demonstrate the utility of a microdroplet emulsions for identifying putative EET-capable bacteria and how this technology can be leveraged in tandem with existing methods.We recently utilized fluorescence from the EET-driven synthesis of a cycloaddition probe, CalFluor488 51 , to assess EET activity from the bacteria S. oneidensis via Cu(I)-catalyzed alkyneazide cycloaddition (CuAAC) 52 . We determined that the rate of copper reduction was directly correlated to EET flux via the well-defined EET-protein pathway in S. oneidensis (the Mtrpathway), and that fluorescence was tied to electron transfer.…”