Rational design of covalent multiheme cytochrome-graphitic carbon dot biohybrids for photo-induced electron transfer

Biohybrid systems can combine inorganic light-harvesting materials and whole-cell biocatalysts to utilize solar energy for the production of chemicals and fuels. Whole-cell biocatalysts have an intrinsic self-repair ability and are able to produce a wide variety of multicarbon chemicals in a sustainable way with metabolic engineering. Current whole-cell biohybrid systems have a yet undefined electron transfer pathway between the light-absorber and metabolic enzymes, limiting rational design. To enable engineering of efficient electron transfer pathways, covalent biohybrids consisting of graphitic nitrogen doped carbon dots (g-N-CDs) and the outer-membrane decaheme protein, MtrC from Shewanella oneidensis MR-1 are developed. MtrC is a subunit of the MtrCAB protein complex, which provides a direct conduit for bidirectional electron exchange across the bacterial outer membrane. The g-N-CDs are functionalized with a maleimide moiety by either carbodiimide chemistry or acyl chloride activation and coupled to a surface-exposed cysteine of a Y657C MtrC mutant. MtrC∼g-N-CD biohybrids are characterized by native and denaturing gel electrophoresis, chromatography, microscopy, and fluorescence lifetime spectroscopy. In the presence of a sacrificial electron donor, visible light irradiation of the MtrC∼g-N-CD biohybrids results in reduced MtrC. The biohybrids may find application in photoinduced transmembrane electron transfer in S. oneidensis MR-1 for chemical synthesis in the future.