Abstract
The low efficiency of extracellular electron transfer (EET) is a major bottleneck for Shewanella oneidensis MR-1 acting as an electroactive biocatalyst in bioelectrochemical systems. Although it is well established that a periplasmic c-type cytochrome (c-Cyt) network plays a critical role in regulating EET efficiency, the understanding of the network in terms of structure and electron transfer activity is obscure and partial. In this work, we attempted to systematically investigate the impacts of the network components on EET in their absence and overproduction individually in microbial fuel cell (MFC). We found that overexpression of c-Cyt CctA leads to accelerated electron transfer between CymA and the Mtr system, which function as the primary quinol oxidase and the outer-membrane (OM) electron hub in EET. In contrast, NapB, FccA, and TsdB in excess severely impaired EET, reducing EET capacity in MFC by more than 50%. Based on the results from both strategies, a series of engineered strains lacking FccA, NapB, and TsdB in combination while overproducing CctA were tested for a maximally optimized c-Cyt network. A strain depleted of all NapB, FccA, and TsdB with CctA overproduction achieved the highest maximum power density in MFCs (436.5 mW/m2), ∼3.62-fold higher than that of wild type (WT). By revealing that optimization of periplasmic c-Cyt composition is a practical strategy for improving EET efficiency, our work underscores the importance in understanding physiological and electrochemical characteristics of c-Cyts involved in EET.
Highlights
Electroactive bacteria capable of extracellular electron transfer (EET) have shown great potential in acting as an electroactive biocatalyst in environmentally friendly bioelectrochemical systems, such as microbial fuel cells (MFCs) and electrolysis cells (Logan and Rabaey, 2012; Shi et al, 2016)
Expression of an additional copy of ccmFGH driven by isopropyl β-D-1-thiogalactoside (IPTG)-inducible promoter Ptac with 0.05 mM IPTG and above enabled wild type (WT) (WT/pccmFGH, grown on TMAO, an electron acceptors (EAs) that supports growth best) to produce more c-type cytochrome (c-Cyt), which can be readily judged by deepened reddish-brown color of cell pellets, a signature feature endowed by c-Cyts (Figure 2A)
The ultimate protein basis for conductivity of microbial EET mediated by both direct contact and indirect electron shuttling via flavin is c-Cyts (Shi et al, 2016; Light et al, 2018; Wang F. et al, 2019; Edwards et al, 2020)
Summary
Electroactive bacteria capable of extracellular electron transfer (EET) have shown great potential in acting as an electroactive biocatalyst in environmentally friendly bioelectrochemical systems, such as microbial fuel cells (MFCs) and electrolysis cells (Logan and Rabaey, 2012; Shi et al, 2016). The periplasmic network is composed of a large number of c-Cyts, which may function as terminal reductases only (i.e., pentaheme nitrite reductase NrfA), electron carriers only [i.e., CctA ( STC)], or both (i.e., fumarate reductase FccA) (Saffarini et al, 2003; Gao et al, 2009; Fonseca et al, 2013). This network mediates electron transport from the quinol oxidases to the Mtr system, an outer-membrane (OM) c-Cyt metal-reducing complex consisting of MtrCAB and OmcA (Shi et al, 2016). Mtr system is responsible for reduction of extracellular electron acceptors (EAs), such as insoluble metal oxides and electrodes, via EET either by the direct interaction between Mtr on the cell surface and EAs or by the indirect approach mediated by flavin electron shuttles (Kotloski and Gralnick, 2013)
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