Single-cell multimodal imaging uncovers energy conversion pathways in biohybrids.

Microbe-semiconductor biohybrids, which integrate microbial enzymatic synthesis with the light-harvesting capabilities of inorganic semiconductors, have emerged as promising solar-to-chemical conversion systems. Improving the electron transport at the nano-bio interface and inside cells is important for boosting conversion efficiencies, yet the underlying mechanism is challenging to study by bulk measurements owing to the heterogeneities of both constituents. Here we develop a generalizable, quantitative multimodal microscopy platform that combines multi-channel optical imaging and photocurrent mapping to probe such biohybrids down to single- to sub-cell/particle levels. We uncover and differentiate the critical roles of different hydrogenases in the lithoautotrophic bacterium Ralstonia eutropha for bioplastic formation, discover this bacterium's surprisingly large nanoampere-level electron-uptake capability, and dissect the cross-membrane electron-transport pathways. This imaging platform, and the associated analytical framework, can uncover electron-transport mechanisms in various types of biohybrid, and potentially offers a means to use and engineer R. eutropha for efficient chemical production coupled with photocatalytic materials.

Thermodynamic Constraints on Electromicrobial Protein Production.

Global consumption of protein is projected to double by the middle of the 21st century. However, protein production is one of the most energy intensive and environmentally damaging parts of the food supply system today. Electromicrobial production technologies that combine renewable electricity and CO2-fixing microbial metabolism could dramatically increase the energy efficiency of commodity chemical production. Here we present a molecular-scale model that sets an upper limit on the performance of any organism performing electromicrobial protein production. We show that engineered microbes that fix CO2 and N2 using reducing equivalents produced by H2-oxidation or extracellular electron uptake could produce amino acids with energy inputs as low as 64 MJ kg-1, approximately one order of magnitude higher than any previous estimate of the efficiency of electromicrobial protein production. This work provides a roadmap for development of engineered microbes that could significantly expand access to proteins produced with a low environmental footprint.

Identification of a pathway for electron uptake in Shewanella oneidensis.

Extracellular electron transfer (EET) could enable electron uptake into microbial metabolism for the synthesis of complex, energy dense organic molecules from CO2 and renewable electricity1-6. Theoretically EET could do this with an efficiency comparable to H2-oxidation7,8 but without the need for a volatile intermediate and the problems it causes for scale up9. However, significant gaps remain in understanding the mechanism and genetics of electron uptake. For example, studies of electron uptake in electroactive microbes have shown a role for the Mtr EET complex in the electroactive microbe Shewanella oneidensis MR-110-14, though there is substantial variation in the magnitude of effect deletion of these genes has depending on the terminal electron acceptor used. This speaks to the potential for previously uncharacterized and/or differentially utilized genes involved in electron uptake. To address this, we screened gene disruption mutants for 3667 genes, representing ≈99% of all nonessential genes, from the S. oneidensis whole genome knockout collection using a redox dye oxidation assay. Confirmation of electron uptake using electrochemical testing allowed us to identify five genes from S. oneidensis that are indispensable for electron uptake from a cathode. Knockout of each gene eliminates extracellular electron uptake, yet in four of the five cases produces no significant defect in electron donation to an anode. This result highlights both distinct electron uptake components and an electronic connection between aerobic and anaerobic electron transport chains that allow electrons from the reversible EET machinery to be coupled to different respiratory processes in S. oneidensis. Homologs to these genes across many different genera suggesting that electron uptake by EET coupled to respiration could be widespread. These gene discoveries provide a foundation for: studying this phenotype in exotic metal-oxidizing microbes, genetic optimization of electron uptake in S. oneidensis; and genetically engineering electron uptake into a highly tractable host like E. coli to complement recent advances in synthetic CO2 fixation15.

Electrical energy storage with engineered biological systems.

The availability of renewable energy technologies is increasing dramatically across the globe thanks to their growing maturity. However, large scale electrical energy storage and retrieval will almost certainly be a required in order to raise the penetration of renewable sources into the grid. No present energy storage technology has the perfect combination of high power and energy density, low financial and environmental cost, lack of site restrictions, long cycle and calendar lifespan, easy materials availability, and fast response time. Engineered electroactive microbes could address many of the limitations of current energy storage technologies by enabling rewired carbon fixation, a process that spatially separates reactions that are normally carried out together in a photosynthetic cell and replaces the least efficient with non-biological equivalents. If successful, this could allow storage of renewable electricity through electrochemical or enzymatic fixation of carbon dioxide and subsequent storage as carbon-based energy storage molecules including hydrocarbons and non-volatile polymers at high efficiency. In this article we compile performance data on biological and non-biological component choices for rewired carbon fixation systems and identify pressing research and engineering challenges.