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ABSTRACT Extracellular iodate reduction by Shewanella spp. contributes to iodide generation in the biogeochemical cycling of iodine. However, there is a disagreement on whether Shewanella spp. use different extracellular electron transfer pathways with dependence on electron donors in iodate reduction. In this study, a series of gene deletion mutants of Shewanella oneidensis MR-1 were created to investigate the roles of dmsEFABGH, mtrCAB, and so4357–so4362 operons in iodate reduction. The iodate-reducing activity of the mutants was tested with lactate, formate, and H2 as the sole electron donors, respectively. In the absence of single-dms gene, iodate reduction efficiency of the mutants was only 12.9%–84.0% with lactate at 24 hours, 22.1%–85.9% with formate at 20 hours, and 19.6%–57.7% with H2 at 42 hours in comparison to complete reduction by the wild type. Progressive inhibition of iodate reduction was observed when the dms homolog from the so4357–so4362 operon was deleted in the single-dms gene mutants. This result revealed complementation of dmsEFABGH by so4357–so4362 at the single-gene level, indicating modularity of the extracellular electron transfer pathway encoded by dmsEFABGH operon. Under the conditions of all electron donors, significant inhibition of iodate reduction and accumulation of H2O2 were detected for ΔmtrCAB. Collectively, these results demonstrated that the dmsEFABGH operon encodes an essential and modular iodate-reducing pathway without electron donor dependence in S. oneidensis MR-1. The mtrCAB operon was involved in H2O2 elimination with all electron donors. The findings in this study improved the understanding of molecular mechanisms underlying extracellular iodate reduction.IMPORTANCEIodine is an essential trace element for human and animals. Recent studies revealed the contribution of microbial extracellular reduction of iodate in biogeochemical cycling of iodine. Multiple reduced substances can be utilized by microorganisms as energy source for iodate reduction. However, varied electron transfer pathways were proposed for iodate reduction with different electron donors in the model strain Shewanella oneidensis MR-1. Here, through a series of gene deletion and iodate reduction experiments, we discovered that the dmsEFABGH operon was essential for iodate reduction with at least three electron donors, including lactate, formate, and H2. The so4357–so4362 operon was first demonstrated to be capable of complementing the function of dmsEFABGH at single-gene level.

Abstract Direct interspecies electron transfer (DIET) may be most important in methanogenic environments, but mechanistic studies of DIET to date have primarily focused on cocultures in which fumarate was the terminal electron acceptor. To better understand DIET with methanogens, the transcriptome of Geobacter metallireducens during DIET‐based growth with G. sulfurreducens reducing fumarate was compared with G. metallireducens grown in coculture with diverse Methanosarcina. The transcriptome of G. metallireducens cocultured with G. sulfurreducens was significantly different from those with Methanosarcina. Furthermore, the transcriptome of G. metallireducens grown with Methanosarcina barkeri, which lacks outer‐surface c‐type cytochromes, differed from those of G. metallireducens cocultured with M. acetivorans or M. subterranea, which have an outer‐surface c‐type cytochrome that serves as an electrical connect for DIET. Differences in G. metallireducens expression patterns for genes involved in extracellular electron transfer were particularly notable. Cocultures with c‐type cytochrome deletion mutant strains, ∆Gmet_0930, ∆Gmet_0557 and ∆Gmet_2896, never became established with G. sulfurreducens but adapted to grow with all three Methanosarcina. Two porin–cytochrome complexes, PccF and PccG, were important for DIET; however, PccG was more important for growth with Methanosarcina. Unlike cocultures with G. sulfurreducens and M. acetivorans, electrically conductive pili were not needed for growth with M. barkeri. Shewanella oneidensis, another electroactive microbe with abundant outer‐surface c‐type cytochromes, did not grow via DIET. The results demonstrate that the presence of outer‐surface c‐type cytochromes does not necessarily confer the capacity for DIET and emphasize the impact of the electron‐accepting partner on the physiology of the electron‐donating DIET partner.

Abstract Purpose c-Type cytochromes are essential for extracellular electron transfer (EET) in electroactive microorganisms. The expression of appropriate c-type cytochromes is an important feature of these microorganisms in response to different extracellular electron acceptors. However, how these diverse c-type cytochromes are tightly regulated is still poorly understood. Methods In this study, we identified the high co-fitness genes that potentially work with different c-type cytochromes by using genome-wide co-fitness analysis. We also constructed and studied the co-fitness networks that composed of c-type cytochromes and the top 20 high co-fitness genes of them. Results We found that high co-fitness genes of c-type cytochromes were enriched in signal transduction processes in Shewanella oneidensis MR-1 cells. We then checked the top 20 co-fitness proteins for each of the 41 c-type cytochromes and identified the corresponding signal proteins for different c-type cytochromes. In particular, through the analysis of the high co-fitness signal protein for CymA, we further confirmed the cooperation between signal proteins and c-type cytochromes and identified a novel signal protein that is putatively involved in the regulation of CymA. In addition, we showed that these signal proteins form two signal transduction modules. Conclusion Taken together, these findings provide novel insights into the coordinated utilization of different c-type cytochromes under diverse conditions.

Electroactive bacteria (EAB) can transfer electrons to exocellular solid acceptors and have widespread applications in the fields of pollutant degradation, biosynthesis, and hydrogen generation. Traditional EAB screening methods can precisely measure the bacterial extracellular electron transfer ability, but they suffer from time-consuming and labor-intensive procedures. The chemiluminescence technique is effective for the rapid detection of hemoglobin but has not been applied for EAB screening. Herein, we utilized a chemiluminescent assay to identify EAB through a multiheme c-type cytochrome (c-Cyt) triggered chemiluminescent reaction. The multiheme c-Cyts-triggered chemiluminescence was conveniently determined by taking photographs with commercial smartphones and cloud-based computing. The acquired image signal shows a significant and linear relationship with the bacterial concentration. Measurement results of the chemiluminescence intensity triggered by different bacterial species with the proposed method are strongly associated with those of the microplate reading method (Pearson's r of 0.9795; P < 0.01) and dissimilatory Fe(III) reduction method (Pearson's r of 0.9628; P < 0.01). Furthermore, this cloud-based and smartphone-assisted chemiluminescent assay is instrument-free and easy to operate, and results can be obtained within 2 min. The findings demonstrate that the proposed method can effectively detect EAB. Therefore, it will be an alternative approach for EAB screening and can have promising applications in microbiology, environmental science, and bioenergy. [ABSTRACT FROM AUTHOR]

During extracellular electron transfer, Geobacter sulfurreducens constitutively expresses the bc‐cytochrome CbcL, yet cells containing only this menaquinone oxidase fail to respire above −0.1 V vs. SHE. By identifying mutations within cbcL that permit growth at higher potentials, we provide evidence that this cytochrome is regulated by redox potential. Strains expressing only CbcLV205A, CbcLV205G, and CbcLF525Y were capable of growth with high potential electron acceptors including Fe(III) citrate, Mn(IV) oxides, and electrodes poised at +0.1 V vs. SHE. Electrochemical characterization of wild type CbcL revealed oxidative inactivation of electron transfer above −0.1 V, while CbcLV205A, CbcLV205G, and CbcLF525Y remained active. Growth yields of CbcLV205A, CbcLV205G, and CbcLF525Y were only 50 % of WT, consistent with CbcL‐dependent electron transfer conserving less energy. These data support the hypothesis that CbcL has evolved to rapidly shut off in response to redox potential, in order to divert electrons to higher yield oxidases that coexist in the Geobacter membrane. [ABSTRACT FROM AUTHOR]

Electron harvesting bacteria are key targets to develop microbial electrosynthesis technologies, which are valid alternatives for the production of value‐added compounds without utilization of fossil fuels. Geobacter sulfurreducens, that is capable of donating and accepting electrons from electrodes, is one of the most promising electroactive bacteria. Its electron transfer mechanisms to electrodes have been progressively elucidated, however the electron harvesting pathways are still poorly understood. Previous studies showed that the periplasmic cytochromes PccH and GSU2515 are overexpressed in current‐consuming G. sulfurreducens biofilms. PccH was characterized, though no putative partners have been identified. In this work, GSU2515 was characterized by complementary biophysical techniques and in silico simulations using the AlphaFold neural network. GSU2515 is a low‐spin monoheme cytochrome with a disordered N‐terminal region and an α‐helical C‐terminal domain harboring the heme group. The cytochrome undergoes a redox‐linked heme axial ligand switch, with Met91 and His94 as distal axial ligands in the reduced and oxidized states, respectively. The reduction potential of the cytochrome is negative and modulated by the pH in the physiological range: −78 mV at pH 6 and −113 mV at pH 7. Such pH‐dependence coupled to the redox‐linked switch of the axial ligand allows the cytochrome to drive a proton‐coupled electron transfer step that is crucial to confer directionality to the respiratory chain. Biomolecular interactions and electron transfer experiments indicated that GSU2515 and PccH form a redox complex. Overall, the data obtained highlight for the first time how periplasmic proteins bridge the electron transfer between the outer and inner membrane in the electron harvesting pathways of G. sulfurreducens. [ABSTRACT FROM AUTHOR]

Mercury nanoparticles are abundant in natural environments. Yet, understanding their contribution to global biogeochemical cycling of mercury remains elusive. Here, we show that microbial transformation of nanoparticulate divalent mercury can be an important source of elemental and methylmercury. Geobacter sulfurreducens PCA, a model bacterium predominant in anoxic environments (e.g., paddy soils), simultaneously reduces and methylates nanoparticulate Hg(II). Moreover, the relative prevalence of these two competing processes and the dominant transformation pathways differ markedly between nanoparticulate Hg(II) and its dissolved and bulk-sized counterparts. Notably, even when intracellular reduction of Hg(II) nanoparticles is constrained by cross-membrane transport (a rate-limiting step that also regulates methylation), the overall Hg(0) formation remains substantial due to extracellular electron transfer. With multiple lines of evidence based on microscopic and electrochemical analyses, gene knockout experiments, and theoretical calculations, we show that nanoparticulate Hg(II) is preferentially associated with c -type cytochromes on cell membranes and has a higher propensity for accepting electrons from the heme groups than adsorbed ionic Hg(II), which explains the surprisingly larger extent of reduction of nanoparticles than dissolved Hg(II) at relatively high mercury loadings. These findings have important implications for the assessment of global mercury budgets as well as the bioavailability of nanominerals and mineral nanoparticles.

Electroactive bacteria have a wide range of applications, including electricity production, bioremediation, and the sensing of toxic compounds. Bacterial biofilm formation is often mediated by the second messenger cyclic guanosine monophosphate (c-di-GMP) synthesized by a diguanylate cyclase (DGC). The role of c-di-GMP in the expression of c-type cytochromes has been previously reported. The aim of this study was to determine the bioelectrogenic activity of Cupriavidus metallidurans strain CH34 pJBpleD*, which possesses a constitutively active DGC that increases c-di-GMP levels. Notably, the heterologous expression of the constitutively active DGC in C. metallidurans strain CH34 pJBpleD* showed a higher biofilm formation and increased the electrical current production up to 560%. In addition, C. metallidurans CH34 pJBpleD* showed increased levels of c-type cytochrome-associated transcripts compared with the wild-type strain CH34. Scanning electron microscopies revealed a denser extracellular matrix with an increased exopolymeric substance content in the CH34 pJBpleD* biofilm on the electrode surface. The results of this study suggest that higher levels of c-di-GMP synthesized by a constitutively active diguanylate cyclase in C. metallidurans strain CH34 pJBpleD* activated the formation of an electroactive biofilm on the electrode, enhancing its exoelectrogenic activity. [ABSTRACT FROM AUTHOR]

Abstract Background Halotolerant Fe (III) oxide reducers affiliated in the family Desulfuromonadaceae are ubiquitous and drive the carbon, nitrogen, sulfur and metal cycles in marine subsurface sediment. Due to their possible application in bioremediation and bioelectrochemical engineering, some of phylogenetically close Desulfuromonas spp. strains have been isolated through enrichment with crystalline Fe (III) oxide and anode. The strains isolated using electron acceptors with distinct redox potentials may have different abilities, for instance, of extracellular electron transport, surface recognition and colonization. The objective of this study was to identify the different genomic signatures between the crystalline Fe (III) oxide-stimulated strain AOP6 and the anode-stimulated strains WTL and DDH964 by comparative genome analysis. Results The AOP6 genome possessed the flagellar biosynthesis gene cluster, as well as diverse and abundant genes involved in chemotaxis sensory systems and c-type cytochromes capable of reduction of electron acceptors with low redox potentials. The WTL and DDH964 genomes lacked the flagellar biosynthesis cluster and exhibited a massive expansion of transposable gene elements that might mediate genome rearrangement, while they were deficient in some of the chemotaxis and cytochrome genes and included the genes for oxygen resistance. Conclusions Our results revealed the genomic signatures distinctive for the ferric iron oxide- and anode-stimulated Desulfuromonas spp. strains. These findings highlighted the different metabolic abilities, such as extracellular electron transfer and environmental stress resistance, of these phylogenetically close bacterial strains, casting light on genome evolution of the subsurface Fe (III) oxide reducers.

Purpose: c-Type cytochromes are essential for extracellular electron transfer (EET) in electroactive microorganisms. The expression of appropriate c-type cytochromes is an important feature of these microorganisms in response to different extracellular electron acceptors. However, how these diverse c-type cytochromes are tightly regulated is still poorly understood. Methods: In this study, we identified the high co-fitness genes that potentially work with different c-type cytochromes by using genome-wide co-fitness analysis. We also constructed and studied the co-fitness networks that composed of c-type cytochromes and the top 20 high co-fitness genes of them. Results: We found that high co-fitness genes of c-type cytochromes were enriched in signal transduction processes in Shewanella oneidensis MR-1 cells. We then checked the top 20 co-fitness proteins for each of the 41 c-type cytochromes and identified the corresponding signal proteins for different c-type cytochromes. In particular, through the analysis of the high co-fitness signal protein for CymA, we further confirmed the cooperation between signal proteins and c-type cytochromes and identified a novel signal protein that is putatively involved in the regulation of CymA. In addition, we showed that these signal proteins form two signal transduction modules. Conclusion: Taken together, these findings provide novel insights into the coordinated utilization of different c-type cytochromes under diverse conditions. [ABSTRACT FROM AUTHOR]

Extracellular iodate reduction by Shewanella spp. contributes to iodide generation in the biogeochemical cycling of iodine. However, there is a disagreement on whether Shewanella spp. use different extracellular electron transfer pathways with dependence on electron donors in iodate reduction. In this study, a series of gene deletion mutants of Shewanella oneidensis MR-1 were created to investigate the roles of dmsEFABGH , mtrCAB, and so4357-so4362 operons in iodate reduction. The iodate-reducing activity of the mutants was tested with lactate, formate, and H 2 as the sole electron donors, respectively. In the absence of single- dms gene, iodate reduction efficiency of the mutants was only 12.9%-84.0% with lactate at 24 hours, 22.1%-85.9% with formate at 20 hours, and 19.6%-57.7% with H 2 at 42 hours in comparison to complete reduction by the wild type. Progressive inhibition of iodate reduction was observed when the dms homolog from the so4357-so4362 operon was deleted in the single- dms gene mutants. This result revealed complementation of dmsEFABGH by so4357-so4362 at the single-gene level, indicating modularity of the extracellular electron transfer pathway encoded by dmsEFABGH operon. Under the conditions of all electron donors, significant inhibition of iodate reduction and accumulation of H 2 O 2 were detected for Δ mtrCAB . Collectively, these results demonstrated that the dmsEFABGH operon encodes an essential and modular iodate-reducing pathway without electron donor dependence in S. oneidensis MR-1. The mtrCAB operon was involved in H 2 O 2 elimination with all electron donors. The findings in this study improved the understanding of molecular mechanisms underlying extracellular iodate reduction.IMPORTANCEIodine is an essential trace element for human and animals. Recent studies revealed the contribution of microbial extracellular reduction of iodate in biogeochemical cycling of iodine. Multiple reduced substances can be utilized by microorganisms as energy source for iodate reduction. However, varied electron transfer pathways were proposed for iodate reduction with different electron donors in the model strain Shewanella oneidensis MR-1. Here, through a series of gene deletion and iodate reduction experiments, we discovered that the dmsEFABGH operon was essential for iodate reduction with at least three electron donors, including lactate, formate, and H 2 . The so4357-so4362 operon was first demonstrated to be capable of complementing the function of dmsEFABGH at single-gene level., Competing Interests: The authors declare no conflict of interest.

Natural microbes employing c‐type cytochromes (c‐Cyts) for extracellular electron transport (EET) were greatly valuable to develop redox‐based bioelectrochemical applications. However, low levels of c‐Cyt expression limited the phylogenetically diverse Gram‐positive species to be used in bioelectrochemical devices. This work reported the remarkable finding that a high level of c‐Cyts was expressed in the cells of the Gram‐positive strain Lysinibacillus varians GY32. c‐Cyts in Lysinibacillus varians GY32 cells were expressed at a level close to those in Shewanella oneidensis. These c‐Cyts were found to cluster in cytoplasmic membrane and periplasmic space, along the length of Lysinibacillus varians GY32 cell. With voltammetry and spectroelectrochemical titration, phenazine methosulfate‐mediated electron transfer between the c‐Cyts and an electrode was proved. These results expanded the accessible natural models of EET and suggested a new microbial platform for developing bioelectrochemical applications. [ABSTRACT FROM AUTHOR]

It is known that the physiology of Methanosarcina species can differ significantly, but the ecological impact of these differences is unclear. We recovered two strains of Methanosarcina from two different ecosystems with a similar enrichment and isolation method. Both strains had the same ability to metabolize organic substrates and participate in direct interspecies electron transfer but also had major physiological differences. Strain DH-1, which was isolated from an anaerobic digester, used H2 as an electron donor. Genome analysis indicated that it lacks an Rnf complex and conserves energy from acetate metabolism via intracellular H2 cycling. In contrast, strain DH-2, a subsurface isolate, lacks hydrogenases required for H2 uptake and cycling and has an Rnf complex for energy conservation when growing on acetate. Further analysis of the genomes of previously described isolates, as well as phylogenetic and metagenomic data on uncultured Methanosarcina in anaerobic digesters and diverse soils and sediments, revealed a physiological dichotomy that corresponded with environment of origin. The physiology of type I Methanosarcina revolves around H2 production and consumption. In contrast, type II Methanosarcina species eschew H2 and have genes for an Rnf complex and the multiheme, membrane-bound c-type cytochrome MmcA, shown to be essential for extracellular electron transfer. The distribution of Methanosarcina species in diverse environments suggests that the type I H2-based physiology is well suited for high-energy environments, like anaerobic digesters, whereas type II Rnf/cytochrome-based physiology is an adaptation to the slower, steady-state carbon and electron fluxes common in organic-poor anaerobic soils and sediments. [ABSTRACT FROM AUTHOR]

Background: Halotolerant Fe (III) oxide reducers affiliated in the family Desulfuromonadaceae are ubiquitous and drive the carbon, nitrogen, sulfur and metal cycles in marine subsurface sediment. Due to their possible application in bioremediation and bioelectrochemical engineering, some of phylogenetically close Desulfuromonas spp. strains have been isolated through enrichment with crystalline Fe (III) oxide and anode. The strains isolated using electron acceptors with distinct redox potentials may have different abilities, for instance, of extracellular electron transport, surface recognition and colonization. The objective of this study was to identify the different genomic signatures between the crystalline Fe (III) oxide-stimulated strain AOP6 and the anode-stimulated strains WTL and DDH964 by comparative genome analysis. Results: The AOP6 genome possessed the flagellar biosynthesis gene cluster, as well as diverse and abundant genes involved in chemotaxis sensory systems and c-type cytochromes capable of reduction of electron acceptors with low redox potentials. The WTL and DDH964 genomes lacked the flagellar biosynthesis cluster and exhibited a massive expansion of transposable gene elements that might mediate genome rearrangement, while they were deficient in some of the chemotaxis and cytochrome genes and included the genes for oxygen resistance. Conclusions: Our results revealed the genomic signatures distinctive for the ferric iron oxide- and anode-stimulated Desulfuromonas spp. strains. These findings highlighted the different metabolic abilities, such as extracellular electron transfer and environmental stress resistance, of these phylogenetically close bacterial strains, casting light on genome evolution of the subsurface Fe (III) oxide reducers. [ABSTRACT FROM AUTHOR]

An outer membrane c-type cytochrome (OmcZ) in Geobacter sulfurreducens is essential for optimal current production in microbial fuel cells. OmcZ exists in two forms, small and large, designated OmcZS and OmcZL, respectively. However, it is still not known how these two structures are formed. A mutant with a disruption of the GSU2075 gene encoding a subtilisin-like serine protease (designated ozpA for the OmcZ protease), which is located downstream of omcZ, produced low currents at a level similar to that of the omcZ-deficient mutant strain. Biochemical analyses revealed that the ozpA mutant accumulated OmcZL and did not produce OmcZS, which is thought to be a mature form that is essential for the extracellular electron transfer to the electrode. A heterologous expression system cell lysate from an Escherichia coli strain producing OzpA cleaved OmcZL and generated OmcZS as the proteolytic product. Among the culture supernatant, loosely bound outer surface, and intracellular protein fractions from wild-type G. sulfurreducens, only the culture supernatant protein fraction showed OmcZL cleavage activity, indicating that the mature form of OmcZ, OmcZS, can be produced outside the cells. These results indicate that OzpA is an essential protease for current production via the maturation of OmcZ, and OmcZS is the key to the extracellular electron transfer to electrodes. This proteolytic maturation of OmcZ is a unique regulation among known c-type cytochromes in G. sulfurreducens. [ABSTRACT FROM AUTHOR]

It has been suggested that a few methanogens are capable of extracellular electron transfers. For instance, Methanosarcina barkeri can directly capture electrons from the coexisting microbial cells of other species. Methanothrix harundinacea and Methanosarcina horonobensis retrieve electrons from Geobacter metallireducens via direct interspecies electron transfer (DIET). Recently, Methanobacterium, designated strain YSL, has been found to grow via DIET in the co-culture with Geobacter metallireducens. Methanosarcina acetivorans can perform anaerobic methane oxidation and respiratory growth relying on Fe(III) reduction through the extracellular electron transfer. Methanosarcina mazei is capable of electromethanogenesis under the conditions where electron-transfer mediators like H2 or formate are limited. The membrane-bound multiheme c-type cytochromes (MHC) and electrically-conductive cellular appendages have been assumed to mediate the extracellular electron transfer in bacteria like Geobacter and Shewanella species. These molecules or structures are rare but have been recently identified in a few methanogens. Here, we review the current state of knowledge for the putative extracellular electron transfers in methanogens and highlight the opportunities and challenges for future research.

It has been suggested that a few methanogens are capable of extracellular electron transfers. For instance, Methanosarcina barkeri can directly capture electrons from the coexisting microbial cells of other species. Methanothrix harundinacea and Methanosarcina horonobensis retrieve electrons from Geobacter metallireducens via direct interspecies electron transfer (DIET). Recently, Methanobacterium , designated strain YSL, has been found to grow via DIET in the co-culture with Geobacter metallireducens. Methanosarcina acetivorans can perform anaerobic methane oxidation and respiratory growth relying on Fe(III) reduction through the extracellular electron transfer. Methanosarcina mazei is capable of electromethanogenesis under the conditions where electron-transfer mediators like H2 or formate are limited. The membrane-bound multiheme c-type cytochromes (MHC) and electrically-conductive cellular appendages have been assumed to mediate the extracellular electron transfer in bacteria like Geobacter and Shewanella species. These molecules or structures are rare but have been recently identified in a few methanogens. Here, we review the current state of knowledge for the putative extracellular electron transfers in methanogens and highlight the opportunities and challenges for future research. [ABSTRACT FROM AUTHOR]

The monoheme outer membrane cytochrome F (OmcF) from Geobacter sulfurreducens plays an important role in Fe(III) reduction and electric current production. The electrochemical characterization of this cytochrome has shown that its redox potential is modulated by the solution pH (redox-Bohr effect) endowing the protein with the necessary properties to couple electron and proton transfer in the physiological range. The analysis of the OmcF structures in the reduced and oxidized states showed that with the exception of the side chain of histidine 47 (His47), all other residues with protonatable side chains are distant from the heme iron and, therefore, are unlikely to affect the redox potential of the protein. The protonatable site at the imidazole ring of His47 is in the close proximity to the heme and, therefore, this residue was suggested as the redox-Bohr center. In the present work, we tested this hypothesis by replacing the His47 with non-protonatable residues (isoleucine – OmcFH47I and phenylalanine – OmcFH47F). The structure of the mutant OmcFH47I was determined by X-ray crystallography to 1.13 Å resolution and showed only minimal changes at the site of the mutation. Both mutants were 15N-labeled and their overall folding was confirmed to be the same as the wild-type by NMR spectroscopy. The pH dependence of the redox potential of the mutants was measured by cyclic voltammetry. Compared to the wild-type protein, the magnitude of the redox-Bohr effect in the mutants was smaller, but not fully abolished, confirming the role of His47 on the pH modulation of OmcF’s redox potential. However, the pH effect on the heme substituents’ NMR chemical shifts suggested that the heme propionate P13 also contributes to the overall redox-Bohr effect in OmcF. In physiological terms, the contribution of two independent acid–base centers to the observed redox-Bohr effect confers OmcF a higher versatility to environmental changes by coupling electron/proton transfer within a wider pH range.

Sutherland et al. recently reported the in vitro inhibition of Helicobacter hepaticus cytochrome c (cyt c) synthase, which may inactivate c-type cytochromes in this and in other pathogenic bacteria. Here, I discuss the impact of this work, which identifies this conserved synthase as a potential antibacterial target. [ABSTRACT FROM AUTHOR]

ABSTRACT Extracellular electron exchange in Methanosarcina species and closely related Archaea plays an important role in the global carbon cycle and enhances the speed and stability of anaerobic digestion by facilitating efficient syntrophic interactions. Here, we grew Methanosarcina acetivorans with methanol provided as the electron donor and the humic analogue, anthraquione-2,6-disulfonate (AQDS), provided as the electron acceptor when methane production was inhibited with bromoethanesulfonate. AQDS was reduced with simultaneous methane production in the absence of bromoethanesulfonate. Transcriptomics revealed that expression of the gene for the transmembrane, multiheme, c-type cytochrome MmcA was higher in AQDS-respiring cells than in cells performing methylotrophic methanogenesis. A strain in which the gene for MmcA was deleted failed to grow via AQDS reduction but grew with the conversion of methanol or acetate to methane, suggesting that MmcA has a specialized role as a conduit for extracellular electron transfer. Enhanced expression of genes for methanol conversion to methyl-coenzyme M and the Rnf complex suggested that methanol is oxidized to carbon dioxide in AQDS-respiring cells through a pathway that is similar to methyl-coenzyme M oxidation in methanogenic cells. However, during AQDS respiration the Rnf complex and reduced methanophenazine probably transfer electrons to MmcA, which functions as the terminal reductase for AQDS reduction. Extracellular electron transfer may enable the survival of methanogens in dynamic environments in which oxidized humic substances and Fe(III) oxides are intermittently available. The availability of tools for genetic manipulation of M. acetivorans makes it an excellent model microbe for evaluating c-type cytochrome-dependent extracellular electron transfer in Archaea. IMPORTANCE The discovery of a methanogen that can conserve energy to support growth solely from the oxidation of organic carbon coupled to the reduction of an extracellular electron acceptor expands the possible environments in which methanogens might thrive. The potential importance of c-type cytochromes for extracellular electron transfer to syntrophic bacterial partners and/or Fe(III) minerals in some Archaea was previously proposed, but these studies with Methanosarcina acetivorans provide the first genetic evidence for cytochrome-based extracellular electron transfer in Archaea. The results suggest parallels with Gram-negative bacteria, such as Shewanella and Geobacter species, in which multiheme outer-surface c-type cytochromes are an essential component for electrical communication with the extracellular environment. M. acetivorans offers an unprecedented opportunity to study mechanisms for energy conservation from the anaerobic oxidation of one-carbon organic compounds coupled to extracellular electron transfer in Archaea with implications not only for methanogens but possibly also for Archaea that anaerobically oxidize methane.

Shewanella oneidensis MR-1 is a platform microorganism for understanding extracellular electron transfer (EET) with a fully sequenced and annotated genome. In comparison to other model microorganisms such as Escherichia coli, the available plasmid parts (such as promoters and replicons) are not sufficient to conveniently and quickly fine-tune the expression of multiple genes in S. oneidensis MR-1. Here, we constructed and characterized a plasmid toolkit that contains a set of expression vectors with a combination of promoters, replicons, antibiotic resistance genes, and an RK2 origin of transfer (oriT) cassette, in which each element can be easily changed by fixed restriction enzyme sites. The expression cassette is also compatible with BioBrick synthetic biology standards. Using green fluorescent protein (GFP) as a reporter, we tested and quantified the strength of promoters. The copy number of different replicons was also measured by real-time quantitative PCR. We further transformed two compatible plasmids with different antibiotic resistance genes into the recombinant S. oneidensis MR-1, enabling control over the expression of two different fluorescent proteins. This plasmid toolkit was further used for overexpression of the MtrCAB porin-c-type cytochrome complex in the S. oneidensis ΔmtrA strain. Tungsten trioxide (WO3) reduction and microbial fuel cell (MFC) assays revealed that the EET efficiency was improved most significantly when MtrCAB was expressed at a moderate level, thus demonstrating the utility of the plasmid toolkit in the EET regulation in S. oneidensis. The plasmid toolkit developed in this study is useful for rapid and convenient fine-tuning of gene expression and enhances the ability to genetically manipulate S. oneidensis MR-1.

BACKGROUND: The c‐type cytochrome of the CymA of Shewanella oneidensis MR‐1 is essential for the anaerobic respiration of Shewanella sp. and transfers electrons from the inner membrane to various terminal electron acceptors, such as soluble redox shuttles and insoluble metal oxides. CymA is believed to be a passage to the outer membrane for dissipating the respiratory electron to the carbon electrode in a microbial fuel cell (MFC) with simultaneous electricity generation. While the deletion and heterologous expression of cymA in Escherichia coli have been studied, there are no reports of the overexpression and its effects on the corresponding bioelectrochemical performance in a MFC. RESULTS: The cymA gene was overexpressed in Shewanella oneidensis MR‐1, and its upregulation was examined under aerobic, anaerobic, and MFC operating conditions by a reverse transcription‐polymerase chain reaction (RT‐PCR). Overexpression of the CymA protein was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE). The MFCs inoculated with the engineered strains of MR‐1 achieved a higher maximum power of 0.13 mW and specific growth rate of 0.087 h−1 than those of the wild type MR‐1 (0.11 mW and 0.043 h−1, respectively). The higher electrochemical activity of the mutant strains demonstrated by cyclic voltammetry and linear sweep voltammetry, indicates that more respiratory electrons can be transferred to the electrodes through overexpression of the cymA gene of MR‐1 in a MFC. CONCLUSION: Overexpression of CymA improves the bioelectrochemical performance of MFCs. This suggests that metabolic engineering of a membrane‐associated redox protein, such as CymA, can further improve electricity generation of MFCs and produce an electrochemically enhanced bioprocess. © 2018 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Electricigens can transfer electrons that produced in intracellular metabolic processes to cellular surface to restore extracellular insoluble electron acceptors (extracellular electron transfer, EET). To uncover the molecular mechanisms underlying EET processes, we integrated transcriptome changes accompanying such processes with molecular network. Firstly, time-series expression datasets for Shewanella oneidensis MR-1 under limited/changed O 2 conditions were obtained from the GEO database, and a total of 336 common differentially expressed genes (DEGs) were identified. Then, we constructed the protein–protein interaction (PPI) network that involved in EET processes from these DEGs. Furthermore, by using centralization analysis and community detection, network analysis of the PPI network was performed. Although the fundamental EET genes are similar to previous studies, important new genes have been discovered. Taking together, our study identified many literature-validated genes critical to EET processes, and also proposed some novel genes that were putatively involved in EET processes. [ABSTRACT FROM AUTHOR]

Bacteria display an array of enzymes to detoxify reactive oxygen species that cause damage to DNA and to other biomolecules leading to cell death. Hydrogen peroxide is one of these species, with endogenous and exogenous sources, such as lactic acid bacteria, oxidative burst of the immune system or chemical reactions at oxic-anoxic interfaces. The enzymes that detoxify hydrogen peroxide will be the focus of this review, with special emphasis on bacterial peroxidases that reduce hydrogen peroxide to water. Bacterial peroxidases are periplasmic cytochromes with either two or three c-type haems, which have been classified as classical and non-classical bacterial peroxidases, respectively. Most of the studies have been focus on the classical bacterial peroxidases, showing the presence of a reductive activation in the presence of calcium ions. Mutagenesis studies have clarified the catalytic mechanism of this enzyme and were used to propose an intramolecular electron transfer pathway, with far less being known about the intermolecular electron transfer that occurs between reduced electron donors and the enzyme. The physiological function of these enzymes was not very clear until it was shown, for the non-classical bacterial peroxidase, that this enzyme is required for the bacteria to use hydrogen peroxide as terminal electron acceptor under anoxic conditions. These non-classical bacterial peroxidases are quinol peroxidases that do not require reductive activation but need calcium ions to attain maximum activity and share similar catalytic intermediates with the classical bacterial peroxidases. [ABSTRACT FROM AUTHOR]

Electron exchange reactions between microbial cells and solid materials, referred to as extracellular electron transfer (EET), have attracted attention in the fields of microbial physiology, microbial ecology, and biotechnology. Studies of model species of iron-reducing, or equivalently, current-generating bacteria such as Geobacter spp. and Shewanella spp. have revealed that redox-active proteins, especially outer membrane c-type cytochromes (OMCs), play a pivotal role in the EET process. Recent (meta)genomic analyses have revealed that diverse microorganisms that have not been demonstrated to have EET ability also harbor OMC-like proteins, indicating that EET via OMCs could be more widely preserved in microorganisms than originally thought. A methanotrophic bacterium Methylococcus capsulatus (Bath) was reported to harbor multiple OMC genes whose expression is elevated by Cu starvation. However, the physiological role of these genes is unknown. Therefore, in this study, we explored whether M. capsulatus (Bath) displays EET abilities via OMCs. In electrochemical analysis, M. capsulatus (Bath) generated anodic current only when electron donors such as formate were available, and could reduce insoluble iron oxides in the presence of electron donor compounds. Furthermore, the current-generating and iron-reducing activities of M. capsulatus (Bath) cells that were cultured in a Cu-deficient medium, which promotes high levels of OMC expression, were higher than those cultured in a Cu-supplemented medium. Anodic current production by the Cu-deficient cells was significantly suppressed by disruption of MCA0421, a highly expressed OMC gene, and by treatment with carbon monoxide (CO) gas (an inhibitor of c-type cytochromes). Our results provide evidence of EET in M. capsulatus (Bath) and demonstrate the pivotal role of OMCs in this process. This study raises the possibility that EET to solid compounds is a novel survival strategy of methanotrophic bacteria.

C-type cytochromes (c-cyts) facilitate microbial extracellular electron transfer and play critical roles in biogeochemical cycling, bioelectricity generation and bioremediation. In this study, a simple and effective method has been developed to detect microbial c-cyts by means of peroxidase mimetic reaction on screen-printed carbon electrode (SPCE). To this end, bacteria cells were immobilized onto the working electrode surface of SPCE by a simple drop casting. After introducing 3,3′,5,5′-tetramethylbenzidine (TMB) solution, microbial c-cyts with peroxidase-like activity catalyze the oxidation of TMB in the presence of hydrogen peroxide. The oxidized TMB was electrochemically determined and the current signal was employed to calculate the c-cyts content. This electrochemical method is highly sensitive for microbial c-cyts with a low detection limit of 40.78 fmol and a wide detection range between 51.70 fmol and 6.64 pmol. Moreover, the proposed technique can be universally expanded to detect c-cyts in other bacteria species such as Fontibacter ferrireducens , Pseudomonas aeruginosa , Comamonas guangdongensis and Escherichia coli . Furthermore, the proposed method confers an in situ facile and quantitative c-cyts detection without any destructive sample preparations, complex electrode modifications and expensive enzyme- or metal particle- based signal amplification. The suggested method advances an intelligent strategy for in situ quantification of microbial c-cyts and consequently holds promising application potential in microbiology and environmental science. [ABSTRACT FROM AUTHOR]

[Display omitted] • A periplasmic protein Cyt c 4 is targeted to play a role in photoelectron transfer. • An ITO-bottom electrochemical reactor records the photoelectron uptake process in situ. • UV–Vis spectroscopy detects 3.35 times higher Cyt c at light than in the dark. • The abundance of Cyt c 4 significantly increases with light intensity enhancement. The excellent performance of biosemiconductors in solar energy conversion and environmental remediation has triggered an explosive research in recent years. However, there is still a lack of knowledge on the mechanisms of extracellular photoelectron uptake in biosemiconductors, especially for the denitrifier-based one. Herein, a model biosemiconductor, Thiobacillus denitrificans -cadmium sulfide (T. denitrificans -CdS), for photoelectrotrophic denitrification was used to investigate the mechanism. Electrochemical characterization demonstrated more redox species generation after exposing T. denitrificans -CdS to light than the controls. Photoelectron uptake process was in situ recorded by an ITO-bottom electrochemical reactor. UV–Vis spectroscopy assured more outer-membrane cytochrome c generation and its concentration was 3.35 times higher in the light than that at dark. A coupled technology of SDS–PAGE, heme staining and LC–MS/MS targeted periplasmic cytochrome c 4 (Cyt c 4) emerging in T. denitrificans -CdS (light). Moreover, the expression of the cyt c 4 gene was significantly (p < 0.05) upregulated under light, and its abundance increased from 1.2 ± 0.3 to 4.7 ± 1.2 folds with light intensity changing from 0.5 to 2.0 mW cm−2, supporting its role in the photoelectron uptake process. This work revealed the photoelectron uptake mechanism in T. denitrificans -CdS, and the results will provide guidance for constructing an efficient biosemiconductor system for nitrate removal and conversion driven by light. [ABSTRACT FROM AUTHOR]

Bacterial extracellular electron transfer (EET) plays a key role in various natural and engineering processes. Outer membrane c-type cytochromes (OMCs) are considered to be essential in bacterial EET. However, most bacteria do not have OMCs but have redox proteins other than OMCs in their extracellular polymeric substances of biofilms. We hypothesized that these extracellular non-cytochrome c proteins (ENCP) could contribute to EET, especially with the facilitation of electron mediators. This study compared the electrode respiring capacity of wild type Shewanella decolorationis S12 and an OMC-deficient mutant. Although the OMC-deficient mutant was incapable in direct electricity generation in normal cultivation, it regained electricity generation capacity (26% of the wide type) with the aid of extracellular electron mediator (riboflavin). Further bioelectrochemistry and X-ray photoelectron spectroscopy analysis suggested that the ENCP, such as proteins with Fe–S cluster, may participate in the falvin-mediated EET. The results highlighted an important and direct role of the ENCP, generated by either electricigens or other microbes, in natural microbial EET process with the facilitation of electron mediators.

The membrane-associated c-type cytochromes ( c-Cyts) have been well known as the key enzymes mediating extracellular electron transfer to terminal electron acceptors, resulting in biogeochemical elemental transformation, contaminant degradation, and nutrient cycling. Although c-Cyts-mediated metal reduction or oxidation have been mainly investigated with the purified proteins of metal reducing/oxidizing bacteria, the in vivo behavior of c-Cyts is still unclear, given the difficulty in measuring the proteins of intact cells. Fortunately, the in situ spectroscopy would be ideal for measuring the reaction kinetics of c-Cyts in intact cells under noninvasive physiological conditions. It can also help the establishment of kinetic/thermodynamic models of extracellular electron transfer processes, which are essential to understand the electron transfer mechanisms at the molecular scale. This review briefly summarizes the current advances in spectral methods for examining the c-Cyts in intact cells of dissimilatory metal reducing bacteria and Fe(II)-oxidizing bacteria. [ABSTRACT FROM AUTHOR]

The highly redundant pathways for extracellular electron transfer in Geobacter sulfurreducens must be simplified for this microorganism to serve as an effective chassis for applications such as the development of sensors and biocomputing. Five homologs of the periplasmic c-type cytochromes, PpcA--E, offer the possibility of multiple routes of electron transfer across the periplasm. The presence of a large number of outer membrane c-type cytochromes allows G. sulfurreducens to adapt to disruption of an electron transfer pathway in the outer membrane. A strain in which genes for all five periplasmic cytochromes, PpcA--E, were deleted did not reduce Fe(III). Introducing ppcA under the control of an IPTG-inducible system in the quintuple deletion strain yielded a strain that reduced Fe(III) only in the presence of IPTG. A strain lacking known major outer membrane cytochromes, OmcB, OmcE, OmcS and OmcT, and putative functional homologs of OmcB, did not reduce Fe(III). Introduction of omcB in this septuple deletion strain restored the ability to reduce Fe(III). These results demonstrate that it is possible to trim redundancy from the extracellular electron transfer pathways in G. sulfurreducens in order to construct strains with defined extracellular electron transfer routes. [ABSTRACT FROM AUTHOR]

A new facultative anaerobic exoelectrogenic strain LZ-1, belonging to Citrobacter freundii , has been isolated. This strain can produce current densities of 843.9 and 865.6 μA cm −2 using citrate or acetate as carbon source in a three-electrode configuration. The electricity generation performance was also analyzed in a dual-chamber MFC system, reaching a maximum power density of 1233 mW m −2 . In addition to acetate and citrate, other carbon sources such as pyruvate, formate, acetate, citrate and fumarate could also be utilized to produce current by strain LZ-1. Data supports the presence of electroactive c -type cytochromes in C. freundii sp. when grown on ITO electrodes, by linking spectroscopy and electrochemistry in situ . Since facultative strains possess many desirable properties compared to anaerobic strains, strain LZ-1 represents a promising exoelectrogenic species in engineering of biological catalysts for microbial electrochemistry. [ABSTRACT FROM AUTHOR]

Shewanella oneidensis MR-1 is capable of extracellular electron transfer (EET) and hence has attracted considerable attention. The EET pathways mainly consist of c-type cytochromes, along with some other proteins involved in electron transfer processes. By whole genome study and protein interactions inquisition, we constructed a large-scale electron transfer network containing 2276 interactions among 454 electron transfer related proteins in S. oneidensis MR-1. Using the k-shell decomposition method, we identified and analyzed distinct parts of the electron transfer network. We found that there was a negative correlation between the ks (k-shell values) and the average DR_100 (disordered regions per 100 amino acids) in every shell, which suggested that disordered regions of proteins played an important role during the formation and extension of the electron transfer network. Furthermore, proteins in the top three shells of the network are mainly located in the cytoplasm and inner membrane; these proteins can be responsible for transfer of electrons into the quinone pool in a wide variety of environmental conditions. In most of the other shells, proteins are broadly located throughout the five cellular compartments (cytoplasm, inner membrane, periplasm, outer membrane and extracellular), which ensures the important EET ability of S. oneidensis MR-1. Specifically, the fourth shell was responsible for EET and the c-type cytochromes in the remaining shells of the electron transfer network were involved in aiding EET. Taken together, these results show that there are distinct functional parts in the electron transfer network of S. oneidensis MR-1, and the EET processes could achieve high efficiency through cooperation through such an electron transfer network.

Front Cover: Single Amino Acid Residues Control Potential-Dependent Inactivation of an Inner Membrane bc-Cytochrome (ChemElectroChem 4/2023) Amino acid substitution, bioelectrochemistry, c-type cytochrome, extracellular electron transfer, Geobacter Keywords: amino acid substitution; bioelectrochemistry; c-type cytochrome; extracellular electron transfer; Geobacter EN amino acid substitution bioelectrochemistry c-type cytochrome extracellular electron transfer Geobacter 1 1 1 02/17/23 20230213 NES 230213 B The Front Cover b illustrates our work identifying a single cytochrome in the I Geobacter i cytoplasmic membrane as the site where electron flow is gated, depending on the redox potential of the acceptor outside the cell. [Extracted from the article]

The use of exoelectrogens in microbial fuel cells (MFCs) has given a wide berth to the addition of artificial electron shuttles/conduits as they have the molecular machinery to transfer the electrons exogenously to the electrode surface or to soluble or insoluble electron acceptors. Exoelectrogens transfer the electrons either directly to the electrode surface (via c-Cyts or pili) and/or mediate them by secreting electron shuttles such as, flavins or pyocyanin. Such microorganisms form electroactive biofilms on the electrode surface. They produce cyclopropane fatty acids and exopolysaccahride matrix to modify surface charge, which also provides favorable anchoring points for the retention of c-type cytochromes (c-Cyts). The longer subunit of PilA plays a vital role in cell attachment in the case of a well-known exoelectrogen Geobacter sulfurreducens during biofilm formation. G. sulfurreducens relies on flavin molecules for mediated electron transfer (MET) during initial biofilm formation and on c-Cyts and pili for the direct electron transfer (DET) during the later phase of biofilm formation. A new protein, cbcl inner membrane multiheme c-Cyt has been revealed in G. sulfurreducens that participates in the electron transfer when electron acceptor with low reduction potential (below 0.1 V) is used in the MFCs. On the other hand, inner membrane c-type cytochrome ImcH is involved in the reduction of electron acceptors exhibiting the potential above 0.1 V. Shewanella oneidensis, another exoelectrogen expresses CheA-3 histidine protein kinase for chemotactic responses to electron acceptors. S. oneidensis do not produce pili and utilizes flavin-cytochrome complexes to regulate the electron transfer to the electrode surfaces. The inherent electron transfer rates can be increased in order to improve the MFC performance. Such strategies as the anode surface modification with nanoparticles, expression of the genes for flavin biosynthesis pathway in the exoelectrogens, and chemical treatment of the microbial membrane have shown to increase the current outputs in the MFCs. This article provides the latest information about the exoelectrogens and molecular drivers involved in extracellular electron transfer (EET) mechanisms, and also summarizes the important characteristics of electroactive biofilms. It also highlights the different approaches that have been employed to facilitate the EET mechanisms and some uncommon exoelectrogens used in the MFCs recently. [ABSTRACT FROM AUTHOR]

The structure of cytochrome c ( GSU3274) designated as PccH from Geobacter sulfurreducens was determined at a resolution of 2.0 Å. PccH is a small (15 kDa) cytochrome containing one c-type heme, found to be essential for the growth of G. sulfurreducens with respect to accepting electrons from graphite electrodes poised at −300 mV versus standard hydrogen electrode. with fumarate as the terminal electron acceptor. The structure of PccH is unique among the monoheme cytochromes described to date. The structural fold of PccH can be described as forming two lobes with the heme sandwiched in a cleft between the two lobes. In addition, PccH has a low reduction potential of −24 mV at pH 7, which is unusual for monoheme cytochromes. Based on difference in structure, together with sequence phylogenetic analysis, we propose that PccH can be regarded as a first characterized example of a new subclass of class I monoheme cytochromes. The low reduction potential of PccH may enable the protein to be redox active at the typically negative potential ranges encountered by G. sulfurreducens. Because PccH is predicted to be located in the periplasm of this bacterium, it could not be involved in the first step of accepting electrons from the electrode but is very likely involved in the downstream electron transport events in the periplasm. [ABSTRACT FROM AUTHOR]

Cytochrome c from the thermophilic bacterium Hydrogenobacter thermophilus is a typical c-type cytochrome which binds heme covalently via two thioether bonds between the two heme vinyl groups and two cysteine thiol groups in a CXXCH sequence motif. This protein was converted to a b-type cytochrome by substitution of the two cysteine residues by alanines (Tomlinson and Ferguson in Proc Natl Acad Sci USA 97:5156-5160, ). To probe the significance of the covalent attachment of the heme in the c-type protein, N relaxation and hydrogen exchange studies have been performed for the wild-type and b-type proteins. The two variants share very similar backbone dynamic properties, both proteins showing high N order parameters in the four main helices, with reduced values in an exposed loop region (residues 18-21), and at the C-terminal residue Lys80. Some subtle changes in chemical shift and hydrogen exchange protection are seen between the wild-type and b-type variant proteins, not only for residues at and neighbouring the mutation sites, but also for some residues in the heme binding pocket. Overall, the results suggest that the main role of the covalent linkages between the heme group and the protein chain must be to increase the stability of the protein. [ABSTRACT FROM AUTHOR]

In this study, an anaerobic batch experiment was conducted to investigate the humus- and Fe(III)-reducing ability of a novel humus-reducing bacterium, Thauera humireducens SgZ-1. Inhibition tests were also performed to explore the electron transport pathways with various electron acceptors. The results indicate that in anaerobic conditions, strain SgZ-1 possesses the ability to reduce a humus analog, humic acids, soluble Fe(III), and Fe(III) oxides. Acetate, propionate, lactate, and pyruvate were suitable electron donors for humus and Fe(III) reduction by strain SgZ-1, while fermentable sugars (glucose and sucrose) were not. UV-visible spectra obtained from intact cells of strain SgZ-1 showed absorption peaks at 420, 522, and 553 nm, characteristic of c-type cytochromes (cyt c). Dithionite-reduced cyt c was reoxidized by Fe-EDTA and HFO (hydrous ferric oxide), which suggests that cyt c within intact cells of strain SgZ-1 has the ability to donate electrons to extracellular Fe(III) species. Inhibition tests revealed that dehydrogenases, quinones, and cytochromes b/ c (cyt b/ c) were involved in reduction of AQS (9, 10-anthraquinone-2-sulfonic acid, humus analog) and oxygen. In contrast, only NADH dehydrogenase was linked to electron transport to HFO, while dehydrogenases and cyt b/ c were found to participate in the reduction of Fe-EDTA. Thus, various different electron transport pathways are employed by strain SgZ-1 for different electron acceptors. The results from this study help in understanding the electron transport processes and environmental responses of the genus Thauera. [ABSTRACT FROM AUTHOR]

The ability to perform the very simple oxidation of two molecules of thiosulfate to tetrathionate is widespread among prokaryotes. Despite the prevalent occurrence of tetrathionate formation and its well documented significance within the sulfur cycle, little is known about the enzymes that catalyze the oxidative condensation of two thiosulfate anions. To fill this gap, the thiosulfate dehydrogenase (TsdA) enzyme from the purple sulfur bacterium Allochromatium vinosum was recombinantly expressed in Escherichia coli, purified and crystallized, and a crystallographic data set was collected. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 79.2, b = 69.9, c = 57.9 Å, β = 129.3°, contained one monomer per asymmetric unit and diffracted to a resolution of 1.98 Å. [ABSTRACT FROM AUTHOR]