Abstract Shewanella oneidensisMR‐1 is a promising chassis organism for microbial electrosynthesis because it has a well‐defined biochemical pathway (the Mtr pathway) that can connect extracellular electrodes to respiratory electron carriers inside the cell. We previously found that the Mtr pathway can be used to transfer electrons from a cathode to intracellular electron carriers and drive reduction reactions. In this work, we hypothesized that native NADH dehydrogenases form an essential link between the Mtr pathway and NADH in the cytoplasm. To test this hypothesis, we compared the ability of various mutant strains to accept electrons from a cathode and transfer them to an NADH‐dependent reaction in the cytoplasm, reduction of acetoin to 2,3‐butanediol. We found that deletion of genes encoding NADH dehydrogenases from the genome blocked electron transfer from a cathode to NADH in the cytoplasm, preventing the conversion of acetoin to 2,3‐butanediol. However, electron transfer to fumarate was not blocked by the gene deletions, indicating that NADH dehydrogenase deletion specifically impacted NADH generation and did not cause a general defect in extracellular electron transfer. Proton motive force (PMF) is linked to the function of the NADH dehydrogenases. We added a protonophore to collapse PMF and observed that it blocked inward electron transfer to acetoin but not fumarate. Together these results indicate a link between the Mtr pathway and intracellular NADH. Future work to optimize microbial electrosynthesis inS. oneidensisMR‐1 should focus on optimizing flux through NADH dehydrogenases.
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The electron transport chain of Shewanella oneidensis MR-1 can operate bidirectionally to enable microbial electrosynthesis
ABSTRACT Extracellular electron transfer is a process by which bacterial cells can exchange electrons with a redox-active material located outside of the cell. InShewanella oneidensis, this process is natively used to facilitate respiration using extracellular electron acceptors such as Fe(III) or an anode. Previously, it was demonstrated that this process can be used to drive the microbial electrosynthesis (MES) of 2,3-butanediol (2,3-BDO) inS. oneidensisexogenously expressing butanediol dehydrogenase (BDH). Electrons taken into the cell from a cathode are used to generate NADH, which in turn is used to reduce acetoin to 2,3-BDO via BDH. However, generating NADH via electron uptake from a cathode is energetically unfavorable, so NADH dehydrogenases couple the reaction to proton motive force. We therefore need to maintain the proton gradient across the membrane to sustain NADH production. This work explores accomplishing this task by bidirectional electron transfer, where electrons provided by the cathode go to both NADH formation and oxygen (O2) reduction by oxidases. We show that oxidases use trace dissolved oxygen in a microaerobic bioelectrical chemical system (BES), and the translocation of protons across the membrane during O2reduction supports 2,3-BDO generation. Interestingly, this process is inhibited by high levels of dissolved oxygen in this system. In an aerated BES, O2molecules react with the strong reductant (cathode) to form reactive oxygen species, resulting in cell death. IMPORTANCEMicrobial electrosynthesis (MES) is increasingly employed for the generation of specialty chemicals, such as biofuels, bioplastics, and cancer therapeutics. For these systems to be viable for industrial scale-up, it is important to understand the energetic requirements of the bacteria to mitigate unnecessary costs. This work demonstrates sustained production of an industrially relevant chemical driven by a cathode. Additionally, it optimizes a previously published system by removing any requirement for phototrophic energy, thereby removing the additional cost of providing a light source. We also demonstrate the severe impact of oxygen intrusion into bioelectrochemical systems, offering insight to future researchers aiming to work in an anaerobic environment. These studies provide insight into both the thermodynamics of electrosynthesis and the importance of the bioelectrochemical systems’ design.
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- Award ID(s):
- 1750785
- PAR ID:
- 10497331
- Editor(s):
- Buan, Nicole R.
- Publisher / Repository:
- American Society for Microbiology
- Date Published:
- Journal Name:
- Applied and Environmental Microbiology
- Volume:
- 90
- Issue:
- 1
- ISSN:
- 0099-2240
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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