*6.2. Microbial Electrosynthesis and Whole-Cell Semi-Artificial Photosynthesis*

Microorganisms are of great interest for electrochemical applications because of their self-reproduce nature and diverse metabolic processes. The possibility to use directly the microorganisms overcomes the need to purify and manipulate the proteins. Many microorganisms are exoelectrogens and can communicate with electrodes through extracellular electron transfer (EET), either (1) directly through membrane-bound cytochromes and conductive filaments (nanowires) or (2) via a soluble redox species, such as flavins, either natively secreted by the microorganism or added as a mediator [202].

#### 6.2.1. Microbial Electrosynthesis

Microbial electrosynthesis is an electricity-driven process which generates chemicals using microorganisms as catalysts [203]. To date, the most well-studied exoelectrogens are *Geobacter* spp. and *S. oneidensis* [204]. They are dissimilatory metal-reducing bacteria that can reduce extracellular metal ions (and bulk electrodes) as part of their anaerobic respiration. These electron transfer steps can be reversed to gain electron from electrode for reductive synthesis, some catalysed by membrane enzymes in the microorganisms (Figure 6) [205,206]. It has been shown that *Geobacter* spp. can reduce nitrate to nitrite [207], fumarate to succinate [207] and proton to hydrogen [208] using electrode as the electron donor. *S. oneidensis* was also shown to electrocatalytically reduce fumarate to succinate [209]. It also has been reported that *S. oneidensis* can catalyse CO<sup>2</sup> reduction into formic acid with electron input from the cathode [210]. The cathodic CO<sup>2</sup> reduction in *S. oneidensis* is associated with the electron uptake through outer-membrane c-type cytochromes [211]. The efficiency of the microbial electrosynthesis is mainly limited by the EET. With the genetic engineering of the microorganisms, the EET efficiency can be improved [212–214]. Advances in synthetic biology allow the rational design of non-natural functions in order to increase the diversity of products obtainable from microbial electrosynthesis [215]. For instance, by engineering genes for an ATP-dependent citrate lyase into *Geobacter sulfurreducens*, the microorganism is able to fix CO<sup>2</sup> through a reverse TCA cycle using an electrode as electron donor [216]. A genetically engineered *S. oneidensis* with heterologous Ehrlich pathway genes was shown to produce isobutanol by supplying electricity [217]. It has been demonstrated that *S. oneidensis* can use electrons supplied by an electrode to reduce O2. This study also showed that the cathodic reaction can reduce NAD<sup>+</sup> via proton-pumping NADH oxidase complex I [218]. With addition of a light-driven proton pump (proteorhodopsin) in the *S. oneidensis* to generate proton-motive force, the electron transferred from cathode to quinone pool can be used to reduce NAD<sup>+</sup> to NADH by native NADH dehydrogenases. This was demonstrated by the reduction of acetoin to 2,3-butanediol via a heterologous butanediol dehydrogenase (Bdh) which is a NADH-dependent enzyme [219].

**Figure 6.** Conceptual model of the bidirectional EET pathways in *S. oneidensis* MR-1. The solid arrows indicate the verified electron flow path, and the dashed arrows indicate the possible electron flow. The blue arrows indicate the outward EET pathway, the red arrows indicate the inward EET pathway, and the green arrows indicate the H<sup>+</sup> flow pathway. (**a**) The outward EET toward extracellular acceptors utilizes the metal-reducing (Mtr) pathway. (**b**) The inward EET toward intracellular terminal reductase involves MtrCBA complex, periplasmic FccA, and inner membrane CymA. The cycle of oxidation (Ox) and reduction (Red) states of self-secreted flavins mediated both outward and inward EET. DMSO, dimethyl sulphoxide; Omc, outer-membrane cytochrome. Reprinted with the permission from ref [220]. Copyright (2020), Taylor & Francis.
