Constraint-Based Model of Shewanella oneidensis MR-1 Metabolism: A Tool for Data Analysis and Hypothesis Generation

Shewanellae are gram-negative facultatively anaerobic metal-reducing bacteria commonly found in chemically (i.e., redox) stratified environments. Occupying such niches requires the ability to rapidly acclimate to changes in electron donor/acceptor type and availability; hence, the ability to compete and thrive in such environments must ultimately be reflected in the organization and utilization of electron transfer networks, as well as central and peripheral carbon metabolism. To understand how Shewanella oneidensis MR-1 utilizes its resources, the metabolic network was reconstructed. The resulting network consists of 774 reactions, 783 genes, and 634 unique metabolites and contains biosynthesis pathways for all cell constituents. Using constraint-based modeling, we investigated aerobic growth of S. oneidensis MR-1 on numerous carbon sources. To achieve this, we (i) used experimental data to formulate a biomass equation and estimate cellular ATP requirements, (ii) developed an approach to identify cycles (such as futile cycles and circulations), (iii) classified how reaction usage affects cellular growth, (iv) predicted cellular biomass yields on different carbon sources and compared model predictions to experimental measurements, and (v) used experimental results to refine metabolic fluxes for growth on lactate. The results revealed that aerobic lactate-grown cells of S. oneidensis MR-1 used less efficient enzymes to couple electron transport to proton motive force generation, and possibly operated at least one futile cycle involving malic enzymes. Several examples are provided whereby model predictions were validated by experimental data, in particular the role of serine hydroxymethyltransferase and glycine cleavage system in the metabolism of one-carbon units, and growth on different sources of carbon and energy. This work illustrates how integration of computational and experimental efforts facilitates the understanding of microbial metabolism at a systems level.     

The utility of Shewanella japonica for microbial fuel cells

Shewanella-containing microbial fuel cells (MFCs) typically use the fresh water wild-type strain Shewanella oneidensis MR-1 due to its metabolic diversity and facultative oxidant tolerance. However, S. oneidensis MR-1 is not capable of metabolizing polysaccharides for extracellular electron transfer. The applicability of Shewanella japonica (an agar-lytic Shewanella strain) for power applications was analyzed using a diverse array of carbon sources for current generation from MFCs, cellular physiological responses at an electrode surface, biofilm formation, and the presence of soluble extracellular mediators for electron transfer to carbon electrodes. Critically, air-exposed S. japonica utilizes biosynthesized extracellular mediators for electron transfer to carbon electrodes with sucrose as the sole carbon source.     

Methods for imaging Shewanella oneidensis MR-1 nanofilaments

Nanofilament production by Shewanella oneidensis MR-1 was evaluated as a function of lifestyle (planktonic vs. sessile) under aerobic and anaerobic conditions using different sample preparation techniques prior to imaging with scanning electron microscopy. Nanofilaments could be imaged on MR-1 cells grown in biofilms or planktonically under both aerobic and anaerobic batch culture conditions after fixation, critical point drying and coating with a conductive metal. Critical point drying was a requirement for imaging nanofilaments attached to planktonically grown MR-1 cells, but not for cells grown in a biofilm. Techniques described in this paper cannot be used to differentiate nanowires from pili or flagella.     

Uridine phosphorylase from Shewanella oneidensis MR-1: Heterological expression, regulation, transcription, and properties

A DNA fragment containing a promoter-operator and structural parts of the uridine phosphorylase gene from Shewanella oneidensis MR-1 was cloned. Cross-heterological expression of the udp genes from Sh. oneidensis MR-1 and Escherichia coli under the control of authentic regulatory regions is shown. The UDP protein accumulates in an active form in the cytoplasmic fraction of cells. The recombinant UDP protein from Sh. oneidensis MR-1 obtained by heterological expression was isolated and characterized. E. coli udp gene promoter activity was observed during heterological expression in Sh. oneidensis MR-1 cells under both aerobic and anaerobic conditions.     

Phage-induced lysis enhances biofilm formation in Shewanella oneidensis MR-1

Shewanella oneidensis MR-1 is capable of forming highly structured surface-attached communities. By DNase I treatment, we demonstrated that extracellular DNA (eDNA) serves as a structural component in all stages of biofilm formation under static and hydrodynamic conditions. We determined whether eDNA is released through cell lysis mediated by the three prophages LambdaSo, MuSo1 and MuSo2 that are harbored in the genome of S. oneidensis MR-1. Mutant analyses and infection studies revealed that all three prophages may individually lead to cell lysis. However, only LambdaSo and MuSo2 form infectious phage particles. Phage release and cell lysis already occur during early stages of static incubation. A mutant devoid of the prophages was significantly less prone to lysis in pure culture. In addition, the phage-less mutant was severely impaired in biofilm formation through all stages of development, and three-dimensional growth occurred independently of eDNA as a structural component. Thus, we suggest that in S. oneidensis MR-1 prophage-mediated lysis results in the release of crucial biofilm-promoting factors, in particular eDNA.     

Effect of NAD+-dependent formate dehydrogenase on anaerobic respiration of Shewanella oneidensis MR-1

An expression plasmid was constructed in order to carry out heterologous expression of the gene of the NAD⁺-dependent formate dehydrogenase (FDH) from methylotrophic bacterium Moraxella sp. in the cells of Shewanella oneidensis MR-1 under aerobic and anaerobic conditions. In both modes of cell cultivation, recombinant FDH activity was revealed in the cell lysate of the transformants. In the medium with la? tate as a carbon source, the rate of anaerobic respiration determined as the rate of conversion of fumarate (the electron acceptor) to succinate was higher in the transformant with recombinant FDH. Anaerobic cultivation of the FDH-containing transformant of S. oneidensis MR-1 in a microbial fuel cell (MFC) revealed increased current density.     

Expression of a Tetraheme Protein, Desulfovibrio vulgaris Miyazaki F Cytochrome c3, in Shewanella oneidensis MR-1

Cytochrome c₃ from Desulfovibrio vulgaris Miyazaki F was successfully expressed in the facultative aerobe Shewanella oneidensis MR-1 under anaerobic, microaerophilic, and aerobic conditions, with yields of 0.3 to 0.5 mg of cytochrome/g of cells. A derivative of the broad-host-range plasmid pRK415 containing the cytochrome c₃ gene from D. vulgaris Miyazaki F was used for transformation of S. oneidensis MR-1, resulting in the production of protein product that was indistinguishable from that produced by D. vulgaris Miyazaki F, except for the presence of one extra alanine residue at the N terminus.     

Hydrogen Metabolism in Shewanella oneidensis MR-1

Shewanella oneidensis MR-1 is a facultative sediment microorganism which uses diverse compounds, such as oxygen and fumarate, as well as insoluble Fe(III) and Mn(IV) as electron acceptors. The electron donor spectrum is more limited and includes metabolic end products of primary fermenting bacteria, such as lactate, formate, and hydrogen. While the utilization of hydrogen as an electron donor has been described previously, we report here the formation of hydrogen from pyruvate under anaerobic, stationary-phase conditions in the absence of an external electron acceptor. Genes for the two S. oneidensis MR-1 hydrogenases, hydA, encoding a periplasmic [Fe-Fe] hydrogenase, and hyaB, encoding a periplasmic [Ni-Fe] hydrogenase, were found to be expressed only under anaerobic conditions during early exponential growth and into stationary-phase growth. Analyses of ΔhydA, ΔhyaB, and ΔhydA ΔhyaB in-frame-deletion mutants indicated that HydA functions primarily as a hydrogen-forming hydrogenase while HyaB has a bifunctional role and represents the dominant hydrogenase activity under the experimental conditions tested. Based on results from physiological and genetic experiments, we propose that hydrogen is formed from pyruvate by multiple parallel pathways, one pathway involving formate as an intermediate, pyruvate-formate lyase, and formate-hydrogen lyase, comprised of HydA hydrogenase and formate dehydrogenase, and a formate-independent pathway involving pyruvate dehydrogenase. A reverse electron transport chain is potentially involved in a formate-hydrogen lyase-independent pathway. While pyruvate does not support a fermentative mode of growth in this microorganism, pyruvate, in the absence of an electron acceptor, increased cell viability in anaerobic, stationary-phase cultures, suggesting a role in the survival of S. oneidensis MR-1 under stationary-phase conditions.     

Microbial reduction of Cr(VI) in the presence of chromate conversion coating constituents

The reduction of Cr(VI) by the metal-reducing bacterium Shewanella oneidensis MR-1 was evaluated, to determine the potential for exploiting Cr(VI) bioreduction as a means of treating chromate conversion coating (CCC) waste streams. Inclusion of Cr(VI) at concentrations ≥1 mM inhibited aerobic growth of S. oneidensis, but that organism was able to reduce Cr(VI) at a concentration of up to 1 mM under anaerobic, nongrowth conditions. S. oneidensis reduced Cr(VI) in the presence of common CCC constituents, with the exception of ferricyanide, when these CCC constituents were included at concentrations typical of CCC waste streams. Ferricyanide inhibited neither aerobic growth nor metabolism under aerobic, nitrate- or iron-reducing conditions, suggesting that the ferricyanide-depended inhibition of Cr(VI) reduction is not due to broad metabolic inhibition, but is specific to Cr(VI) reduction. Results indicate that under some conditions, the activities of metal-reducing bacteria, such as S. oneidensis, could be exploited for the removal of Cr(VI) from CCC waste streams under appropriate conditions.     

Shewanella oneidensis MR-1 Msh pilin proteins are involved in extracellular electron transfer in microbial fuel cells

Shewanella is a microbial genus that can oxidize lactate for the reduction of insoluble electron acceptors. This reduction is possible by either direct (cell-surface interaction, nanowires) or indirect (soluble redox mediators) mechanisms. However, the actual molecular identification of a nanowire has not been determined. Through mutational studies, Shewanella oneidensis MR-1 was analyzed for its ability to transfer electrons to an electrode after deletion of the structural pilin genes (ΔmshA-D) or the entire biosynthetic expression system (ΔmshH-Q) of one of its pilin complexes (Msh type IV pilus gene locus). The complete removal of the Msh complex (ΔmshH-Q) significantly decreased the current generated from a fuel cell compared to MR-1. However, the mutant with only extracellular Msh structural proteins removed (ΔmshA-D) was able to generate 80% of the current compared to MR-1. Thus, the intracellular and membrane bound Msh biogenesis complex is a pathway for extracellular electron transfer in S. oneidensis MR-1.     

Substrate-Level Phosphorylation Is the Primary Source of Energy Conservation during Anaerobic Respiration of Shewanella oneidensis Strain MR-1

It is well established that respiratory organisms use proton motive force to produce ATP via F-type ATP synthase aerobically and that this process may reverse during anaerobiosis to produce proton motive force. Here, we show that Shewanella oneidensis strain MR-1, a nonfermentative, facultative anaerobe known to respire exogenous electron acceptors, generates ATP primarily from substrate-level phosphorylation under anaerobic conditions. Mutant strains lacking ackA (SO2915) and pta (SO2916), genes required for acetate production and a significant portion of substrate-level ATP produced anaerobically, were tested for growth. These mutant strains were unable to grow anaerobically with lactate and fumarate as the electron acceptor, consistent with substrate-level phosphorylation yielding a significant amount of ATP. Mutant strains lacking ackA and pta were also shown to grow slowly using N-acetylglucosamine as the carbon source and fumarate as the electron acceptor, consistent with some ATP generation deriving from the Entner-Doudoroff pathway with this substrate. A deletion strain lacking the sole F-type ATP synthase (SO4746 to SO4754) demonstrated enhanced growth on N-acetylglucosamine and a minor defect with lactate under anaerobic conditions. ATP synthase mutants grown anaerobically on lactate while expressing proteorhodopsin, a light-dependent proton pump, exhibited restored growth when exposed to light, consistent with a proton-pumping role for ATP synthase under anaerobic conditions. Although S. oneidensis requires external electron acceptors to balance redox reactions and is not fermentative, we find that substrate-level phosphorylation is its primary anaerobic energy conservation strategy. Phenotypic characterization of an ackA deletion in Shewanella sp. strain MR-4 and genomic analysis of other sequenced strains suggest that this strategy is a common feature of Shewanella.Shewanella oneidensis strain MR-1 is a nonfermentative, facultative anaerobe which respires various substrates, including oxygen, soluble metals, insoluble iron and manganese oxide minerals, electrodes, and organic compounds (8, 12, 18, 22). Other bacteria with the ability to respire electrodes and oxide minerals, such as Geobacter and Geothrix, oxidize acetate to carbon dioxide (4, 7, 9), consistent with these organisms generating ATP primarily from oxidative phosphorylation rather than substrate-level phosphorylation. Yet, an examination of metabolic end products and a variety of central metabolism and flux analyses of MR-1 show that acetate is the major product under anaerobic conditions (18, 27, 29, 31). The general anaerobic metabolism model for MR-1, as depicted in Fig. ​Fig.1,1, has key features of glycolysis via the Entner-Doudoroff pathway as well as acetyl coenzyme A (acetyl-CoA) flux toward acetate anaerobically via phosphate acetyltransferase (Pta) and acetate kinase (AckA) (27, 29, 31). High-performance liquid chromatography (HPLC) studies in our lab and others have shown that pyruvate may be excreted during lactate utilization both aerobically and anaerobically (30, 31), and MR-1 has not been shown to maintain significant flux through the tricarboxylic acid (TCA) cycle under anaerobic conditions (31).Open in a separate windowFIG. 1.Simplified model of S. oneidensis central metabolism. Entner-Doudoroff glycolysis yields two molecules of pyruvate. Under aerobic conditions, pyruvate facilitates the reduction of NAD⁺ to NADH before being completely oxidized to carbon dioxide in the TCA cycle. Anaerobically, pyruvate oxidation to acetyl-CoA yields formate before the pyruvate is converted to acetate. Formate is subsequently oxidized to carbon dioxide. Reactions catalyzed by acetate kinase and phosphate acetyltransferase are denoted AckA and Pta, respectively. QH₂ is reduced quinone. The model is based on several references (5, 24, 29, 31, 36).Characterization studies of proton motive force (PMF) in MR-1 have not definitively determined whether the source of anaerobic proton pumping or translocation is electron transport, ATP synthase, or metabolite transport (13, 19). Myers et al. demonstrated that anaerobic MR-1 cells starved of electron acceptor generate PMF in response to fumarate addition (19). However, the directionality of the ATP synthase (i.e., generation of ATP or ATPase to pump protons) was not characterized. Previous work has confirmed that proteorhodopsin (PR), a light-dependent, proton-pumping integral membrane protein, can be used in MR-1 to supplement PMF (13). However, the observed increase in PMF in wild-type cells expressing PR did not result in higher optical densities (ODs) or in a higher growth rate. Though all known bacteria depend on PMF, whether MR-1 uses that PMF for ATP production or uses ATP to help generate PMF under anaerobic conditions has yet to be determined.To examine ATP production in MR-1, growth on carbon sources that offer various amounts of substrate-level-derived ATP and reducing equivalents (NADH, formate, or quinones) in their oxidation was characterized. Two carbon sources entering central metabolism at different locations are N-acetylglucosamine (NAG) and lactate, which enter before and after glycolysis, respectively (Fig. ​(Fig.1)1) (24, 36). Both are oxidized to acetate and carbon dioxide anaerobically, though lactate yields one ATP and two reducing equivalents per molecule, while NAG yields three ATPs and four reducing equivalents per molecule. The differences in ATP yields derived from utilization of NAG versus lactate, combined with modification of those yields through gene deletions, allowed for characterization of ATP production in MR-1.The goal of this work was to elucidate the primary source of ATP generation under anaerobic conditions in MR-1. Data presented here support a model of anaerobic metabolism where substrate-level phosphorylation is the primary mechanism for ATP generation and where some amount of the ATP pool is used to generate PMF. Paradoxically, the most diverse respiratory organism characterized to date (8, 12, 22) does not generate ATP from electron transport reactions and PMF. Our finding highlights a critical difference in metabolic strategies between Shewanella and other organisms that are able to reduce insoluble substrates, such as Geobacter and Geothrix.     

Shewanella oneidensis MR-1 Fluxome under Various Oxygen Conditions

The central metabolic fluxes of Shewanella oneidensis MR-1 were examined under carbon-limited (aerobic) and oxygen-limited (microaerobic) chemostat conditions, using ¹³C-labeled lactate as the sole carbon source. The carbon labeling patterns of key amino acids in biomass were probed using both gas chromatography-mass spectrometry (GC-MS) and ¹³C nuclear magnetic resonance (NMR). Based on the genome annotation, a metabolic pathway model was constructed to quantify the central metabolic flux distributions. The model showed that the tricarboxylic acid (TCA) cycle is the major carbon metabolism route under both conditions. The Entner-Doudoroff and pentose phosphate pathways were utilized primarily for biomass synthesis (with a flux below 5% of the lactate uptake rate). The anaplerotic reactions (pyruvate to malate and oxaloacetate to phosphoenolpyruvate) and the glyoxylate shunt were active. Under carbon-limited conditions, a substantial amount (9% of the lactate uptake rate) of carbon entered the highly reversible serine metabolic pathway. Under microaerobic conditions, fluxes through the TCA cycle decreased and acetate production increased compared to what was found for carbon-limited conditions, and the flux from glyoxylate to glycine (serine-glyoxylate aminotransferase) became measurable. Although the flux distributions under aerobic, microaerobic, and shake flask culture conditions were different, the relative flux ratios for some central metabolic reactions did not differ significantly (in particular, between the shake flask and aerobic-chemostat groups). Hence, the central metabolism of S. oneidensis appears to be robust to environmental changes. Our study also demonstrates the merit of coupling GC-MS with ¹³C NMR for metabolic flux analysis to reduce the use of ¹³C-labeled substrates and to obtain more-accurate flux values.     

Developing a base-editing system to expand the carbon source utilization spectra of Shewanella oneidensis MR-1 for enhanced pollutant degradation

Shewanella oneidensis MR-1, a model strain of exoelectrogenic bacteria (EEB), plays a key role in environmental bioremediation and bioelectrochemical systems because of its unique respiration capacity. However, only a narrow range of substrates can be utilized by S. oneidensis MR-1 as carbon sources, resulting in its limited applications. In this study, a rapid, highly efficient, and easily manipulated base-editing system pCBEso was developed by fusing a Cas9 nickase (Cas9n (D10A)) with the cytidine deaminase rAPOBEC1 in S. oneidensis MR-1. The C-to-T conversion of suitable C within the base-editing window could be readily and efficiently achieved by the pCBEso system without requiring double-strand break or repair templates. Moreover, double-locus simultaneous editing was successfully accomplished with an efficiency of 87.5%. With this tool, the key genes involving in N-acetylglucosamine (GlcNAc) or glucose metabolism in S. oneidensis MR-1 were identified. Furthermore, an engineered strain with expanded carbon source utilization spectra was constructed and exhibited a higher degradation rate for multiple organic pollutants (i.e., azo dyes and organoarsenic compounds) than the wild-type when glucose or GlcNAc was used as the sole carbon source. Such a base-editing system could be readily applied to other EEB. This study not only enhances the substrate utilization and pollutant degradation capacities of S. oneidensis MR-1 but also accelerates the robust construction of engineered strains for environmental bioremediation.     

A Biochemical Approach to Study the Role of the Terminal Oxidases in Aerobic Respiration in Shewanella oneidensis MR-1

The genome of the facultative anaerobic γ-proteobacterium Shewanella oneidensis MR-1 encodes for three terminal oxidases: a bd-type quinol oxidase and two heme-copper oxidases, a A-type cytochrome c oxidase and a cbb ₃-type oxidase. In this study, we used a biochemical approach and directly measured oxidase activities coupled to mass-spectrometry analysis to investigate the physiological role of the three terminal oxidases under aerobic and microaerobic conditions. Our data revealed that the cbb ₃-type oxidase is the major terminal oxidase under aerobic conditions while both cbb ₃-type and bd-type oxidases are involved in respiration at low-O₂ tensions. On the contrary, the low O₂-affinity A-type cytochrome c oxidase was not detected in our experimental conditions even under aerobic conditions and would therefore not be required for aerobic respiration in S. oneidensis MR-1. In addition, the deduced amino acid sequence suggests that the A-type cytochrome c oxidase is a ccaa ₃-type oxidase since an uncommon extra-C terminal domain contains two c-type heme binding motifs. The particularity of the aerobic respiratory pathway and the physiological implication of the presence of a ccaa ₃-type oxidase in S. oneidensis MR-1 are discussed.     

Mutants of an electrogenic bacterium <Emphasis Type="Italic">Shewanella oneidensis</Emphasis> MR-1 with increased reducing activity

The mutants of Shewanella oneidensis MR-1 resistant to fosfomycin, a toxic analogue of phosphoenolpyruvate, were obtained. The mutants exhibited increased reducing activity and higher rates of lactate utilization. A correlation was shown between the rates of metabolism of oxidized substrates and the rate of reduction of methylene blue, a mediator of electron transport. The mutants of S. oneidensis MR-1 may be used in microbial fuel cells for intensification of energy production from organic compounds.     

Decolorization and detoxification of a sulfonated triphenylmethane dye aniline blue by Shewanella oneidensis MR-1 under anaerobic conditions

In this work, the extracellular decolorization of aniline blue, a sulfonated triphenylmethane dye, by Shewanella oneidensis MR-1 was confirmed. S. oneidensis MR-1 showed a high capacity for decolorizing aniline blue even at a concentration of up to 1,000 mg/l under anaerobic conditions. Maximum decolorization efficiency appeared at pH?7.0 and 30 °C. Lactate was a better candidate of electron donor for the decolorization of aniline blue. The addition of nitrate, hydrous ferric oxide, or trimethylamine N-oxide all could cause a significant decline of decolorization efficiency. The Mtr respiratory pathway was found to be involved into the decolorization of aniline blue by S. oneidensis MR-1. The toxicity evaluation through phytotoxicity and genotoxicity showed that S. oneidensis MR-1 could decrease the toxicity of aniline blue during the decolorization process. Thus, this work may facilitate a better understanding on the degradation mechanisms of the triphenylmethane dyes by Shewanella and is beneficial to their application in bioremediation.     

Current Production and Metal Oxide Reduction by Shewanella oneidensis MR-1 Wild Type and Mutants

Shewanella oneidensis MR-1 is a gram-negative facultative anaerobe capable of utilizing a broad range of electron acceptors, including several solid substrates. S. oneidensis MR-1 can reduce Mn(IV) and Fe(III) oxides and can produce current in microbial fuel cells. The mechanisms that are employed by S. oneidensis MR-1 to execute these processes have not yet been fully elucidated. Several different S. oneidensis MR-1 deletion mutants were generated and tested for current production and metal oxide reduction. The results showed that a few key cytochromes play a role in all of the processes but that their degrees of participation in each process are very different. Overall, these data suggest a very complex picture of electron transfer to solid and soluble substrates by S. oneidensis MR-1.     

Selenite and Tellurite Reduction by Shewanella oneidensis

Shewanella oneidensis MR-1 reduces selenite and tellurite preferentially under anaerobic conditions. The Se(0) and Te(0) deposits are located extracellularly and intracellularly, respectively. This difference in localization and the distinct effect of some inhibitors and electron acceptors on these reduction processes are taken as evidence of two independent pathways.     

A derivative of the menaquinone precursor 1,4-dihydroxy-2-naphthoate is involved in the reductive transformation of carbon tetrachloride by aerobically grown<Emphasis Type="Italic"> Shewanella oneidensis</Emphasis> MR-1

Transformation of carbon tetrachloride (CT) by Shewanella oneidensis MR-1 has been proposed to involve the anaerobic respiratory-chain component menaquinone. To investigate this hypothesis a series of menaquinone mutants were constructed. The menF mutant is blocked at the start of the menaquinone biosynthetic pathway. The menB, menA and menG mutants are all blocked towards the end of the pathway, being unable to produce 1,4-dihydroxy-2-naphthoic acid (DHNA), demethyl-menaquinone and menaquinone , respectively. Aerobically grown mutants unable to produce the menaquinone precursor DHNA (menF and menB mutants) showed a distinctly different CT transformation profile than mutants able to produce DHNA but unable to produce menaquinone (menA and menG mutants). While DHNA did not reduce CT in an abiotic assay, the addition of DHNA to the menF and menB mutants restored normal CT transformation activity. We conclude that a derivative of DHNA, that is distinct from menaquinone, is involved in the reduction of CT by aerobically grown S. oneidensis MR-1. When cells were grown anaerobically with trimethylamine-N-oxide as the terminal electron acceptor, all the menaquinone mutants showed wild-type levels of CT reduction. We conclude that S. oneidensis MR-1 produces two different factors capable of dehalogenating CT. The factor produced under anaerobic growth conditions is not a product of the menaquinone biosynthetic pathway.     

Intensification of bioelectricity generation in microbial fuel cells using Shewanella oneidensis MR-1 mutants with increased reducing activity

Electrogenicity of Shewanella oneidensis MR-1 mutants FRS1 and FRB1 with reducing activity 30–40% higher than in the original strain was studied in various microbial fuel cells (MFC) developed in the course of the work. The voltage and current density developed by the mutants were 1.7 times higher than in the case of S. oneidensis MR-1. A correlation was found between reducing activity of the cells and the voltage and current density developed in MFC. The possibility for enhanced bioelectricity production in MFC by genetic modification of S. oneidensis MR-1 was demonstrated.