3-Hydroxypropionic acid (3-HP) is an important platform chemical which can be produced biologically from glycerol. Klebsiella pneumoniae is an ideal biocatalyst for 3-HP because it can grow well on glycerol and naturally synthesize the essential coenzyme B12. On the other hand, if higher yields and titers of 3-HP are to be achieved, the sustained regeneration of NAD+ under anaerobic conditions, where coenzyme B12 is synthesized sustainably, is required. In this study, recombinant K. pneumoniae L17 overexpressing aldehyde dehydrogenase (AldH) was developed and cultured in a bioelectrochemical system (BES) with the application of an electrical potential to the anode using a chronoamperometric method (+0.5 V vs. Ag/AgCl). The BES operation resulted in 1.7-fold enhancement of 3-HP production compared to the control without the applied potential. The intracellular NADH/NAD+ ratio was significantly lower when the L17 cells were grown under an electric potential. The interaction between the electrode and overexpressed AldH was enhanced by electron shuttling mediated by HNQ (2-hydroxy-1,4-naphthoquinone). Enhanced 3-HP production by the BES was achieved using recombinant K. pneumoniae L17. The quinone-based electron transference between the electrode and L17 was investigated by respiratory uncoupler experiments. This study provides a novel strategy to control the intracellular redox states to enhance the yield and titer of 3-HP production as well as other bioconversion processes.

Mixed microbial culture (MMC) electrofermentation (EF) represents a promising tool to drive metabolic pathways toward the production of a specific compound. Here, the MMC-EF process has been exploited to obtain butyric acid in simplified membrane-less reactors operated by applying a difference of potential between two low-cost graphite electrodes. Ten values of voltage difference, from −0.60 V to −1.5 V, have been tested and compared with the experiment under open circuit potential (OCP). In all the tested conditions, an enhancement in the production rate of butyric acid (from a synthetic mixture of glucose, acetate, and ethanol) was observed, ranging from 1.3- to 2.7-fold relative to the OCP. Smaller enhancements in the production rate resulted in higher values of the calculated specific energy consumption. However, at all applied voltages, a low flow of current was detected in the one-chamber reactors, accounting for an average value of approximately −100 µA. These results hold a substantial potential with respect to the scalability of the electrofermentation technology, since they pinpoint the possibility to control MMC-based bioprocesses by simply inserting polarized electrodes into traditional fermenters.

The electrogenic biofilm and the bio-electrode interface are the key biocatalytic components in bioelectrochemical systems (BES) and can have a large impact on cell performance. This study used four different anodic carbons to investigate electrogenic biofilm development to determine the influence of charge accumulation and biofilm growth on system performance and how biofilm structure may mitigate against pH perturbations. Power production was highest (1.40 W/m³) using carbon felt, but significant power was also produced when felt carbon was open-circuit acclimated in a control reactor (0.95 W/m³). The influence of carbon material on electrogenic biofilm development was determined by measuring the level of biofilm growth, using sequencing to identify the microbial populations and confocal microscopy to understand the spatial locations of key microbial groups. Geobacter spp. were found to be enriched in closed-circuit operation and these were in close association with the carbon anode, but these were not observed in the open-circuit controls. Electrochemical analysis also demonstrated that the highest mid-point anode potentials were close to values reported for cytochromes from Geobacter sulfurreductans. Biofilm development was greatest in felt anodes (closed-circuit acclimated 1209 ng/μL DNA), and this facilitated the highest pseudo-capacitive values due to the presence of redox-active species, and this was associated with higher levels of power production and also served to mitigate against the effects of low-pH operation. Supporting carbon anode structures are key to electrogenic biofilm development and associated system performance and are also capable of protecting electrochemically active bacteria from the effects of environmental perturbations.