Engineering outer-membrane electron transfer proteins for modular biotic-abiotic interfacing with living cells

The electrogenic bacterium Shewanella oneidensis utilises the MtrCAB protein complex to transfer electrons across the outer membrane, forming a biotic-abiotic interface that can be harnessed for energy generation, biosensing, microbial electrosynthesis and cell signalling. Within this complex, the decaheme cytochromes MtrC and MtrA are arranged on the extracellular and periplasmic sides of the outer-membrane β-barrel protein MtrB, respectively, enabling electron transfer between the cell interior and the extracellular environment. Extracellular electron transfer (EET) through MtrCAB supports anaerobic respiration by passing electrons to extracellular acceptors, while extracellular electron uptake (EEU) imports electrons from extracellular donors to support aerobic metabolism. In this thesis, I demonstrate that the C-terminus of MtrC can be engineered to enable selective bioconjugation to materials, which improved energy generation by 30% relative to an unmodified control. A modifiable MtrC offers a modular platform for interfacing living cells with electronic materials, such as electrodes, conductive synthetic biofilms, and redox enzymes for biosensing or biocatalysis. Furthermore, I demonstrate that the N-terminus of MtrA can be modified without disrupting EET. Fusion of a P450 heme domain to MtrA facilitated microbial electrosynthesis driven by EEU from an extracellular cathode. This intracellular electrosynthesis strategy offers advantages over conventional in vitro redox biocatalysis by continuously regenerating the redox protein within the cell and reducing dependence on externally supplied redox cofactors or intermediates. Together, these results establish engineered MtrC and MtrA as modular entry points for constructing multifunctional protein-based biotic-abiotic interfaces and provide a foundation for future electronic control of intracellular processes.

Grantor dc:publisher

UNSW, Sydney

Year dc:date

2026

Author dc:creator

• Kelly, Alexander ; https://orcid.org/0009-0004-3054-2166

• Biotic-Abiotic Interfaces
• Shewanella oneidensis
• Escherichia coli
• Microbial Fuel Cell
• Microbial Electrolysis Cell
• Microbial Electrosynthesis
• SpyTag/SpyCatcher
• MtrC
• MtrA
• GrBP5
• MFC
• MEC
• Protein engineering
• anzsrc-for: 3101 Biochemistry and cell biology
• anzsrc-for: 3107 Microbiology

Statement dc:rights

• open access
• CC BY 4.0
• free_to_read

Licence

• https://purl.org/coar/access_right/c_abf2
• https://creativecommons.org/licenses/by/4.0/

Language dc:language

en

Identifier

https://doi.org/10.26190/unsworks/32439

OAI identifier oai:identifier

oai:unsworks.library.unsw.edu.au:1959.4/108135