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1.
Nature ; 597(7876): 430-434, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34471289

RESUMO

Extracellular electron transfer by Geobacter species through surface appendages known as microbial nanowires1 is important in a range of globally important environmental phenomena2, as well as for applications in bio-remediation, bioenergy, biofuels and bioelectronics. Since 2005, these nanowires have been thought to be type 4 pili composed solely of the PilA-N protein1. However, previous structural analyses have demonstrated that, during extracellular electron transfer, cells do not produce pili but rather nanowires made up of the cytochromes OmcS2,3 and OmcZ4. Here we show that Geobacter sulfurreducens binds PilA-N to PilA-C to assemble heterodimeric pili, which remain periplasmic under nanowire-producing conditions that require extracellular electron transfer5. Cryo-electron microscopy revealed that C-terminal residues of PilA-N stabilize its copolymerization with PilA-C (to form PilA-N-C) through electrostatic and hydrophobic interactions that position PilA-C along the outer surface of the filament. PilA-N-C filaments lack π-stacking of aromatic side chains and show a conductivity that is 20,000-fold lower than that of OmcZ nanowires. In contrast with surface-displayed type 4 pili, PilA-N-C filaments show structure, function and localization akin to those of type 2 secretion pseudopili6. The secretion of OmcS and OmcZ nanowires is lost when pilA-N is deleted and restored when PilA-N-C filaments are reconstituted. The substitution of pilA-N with the type 4 pili of other microorganisms also causes a loss of secretion of OmcZ nanowires. As all major phyla of prokaryotes use systems similar to type 4 pili, this nanowire translocation machinery may have a widespread effect in identifying the evolution and prevalence of diverse electron-transferring microorganisms and in determining nanowire assembly architecture for designing synthetic protein nanowires.


Assuntos
Fímbrias Bacterianas/química , Fímbrias Bacterianas/metabolismo , Geobacter , Nanofios , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Biopolímeros , Condutividade Elétrica , Proteínas de Fímbrias/química , Proteínas de Fímbrias/metabolismo , Geobacter/citologia , Geobacter/metabolismo , Multimerização Proteica
2.
Artigo em Inglês | MEDLINE | ID: mdl-34444543

RESUMO

This paper analyzed the degradation pathways of petroleum hydrocarbon degradation bacteria, screened the main degradation pathways, and found the petroleum hydrocarbon degradation enzymes corresponding to each step of the degradation pathway. Through the Copeland method, the best inoculation program of petroleum hydrocarbon degradation bacteria in a polluted site was selected as follows: single oxygenation path was dominated by Streptomyces avermitilis, hydroxylation path was dominated by Methylosinus trichosporium OB3b, secondary oxygenation path was dominated by Pseudomonas aeruginosa, secondary hydroxylation path was dominated by Methylococcus capsulatus, double oxygenation path was dominated by Acinetobacter baylyi ADP1, hydrolysis path was dominated by Rhodococcus erythropolis, and CoA path was dominated by Geobacter metallireducens GS-15 to repair petroleum hydrocarbon contaminated sites. The Copeland method score for this solution is 22, which is the highest among the 375 solutions designed in this paper, indicating that it has the best degradation effect. Meanwhile, we verified its effect by the Cdocker method, and the Cdocker energy of this solution is -285.811 kcal/mol, which has the highest absolute value. Among the inoculation programs of the top 13 petroleum hydrocarbon degradation bacteria, the effect of the best inoculation program of petroleum hydrocarbon degradation bacteria was 18% higher than that of the 13th group, verifying that this solution has the best overall degradation effect. The inoculation program of petroleum hydrocarbon degradation bacteria designed in this paper considered the main pathways of petroleum hydrocarbon pollutant degradation, especially highlighting the degradability of petroleum hydrocarbon intermediate degradation products, and enriching the theoretical program of microbial remediation of petroleum hydrocarbon contaminated sites.


Assuntos
Petróleo , Rhodococcus , Poluentes do Solo , Acinetobacter , Biodegradação Ambiental , Geobacter , Hidrocarbonetos , Solo , Microbiologia do Solo , Poluentes do Solo/análise , Streptomyces
3.
Water Res ; 202: 117490, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34364064

RESUMO

Geobacter, as a typical electroactive microorganism, is the "engine" of interspecies electron transfer (IET) between microorganisms. However, it does not have a dominant position in all natural environments. It is not known what performs a similar function as Geobacter in coastal zones. Metagenomic and metatranscriptomic analysis revealed that Desulfovibrio and Methanobacterium species were the most abundant in electrochemically active aggregates. Metatranscriptomic analysis showed that Desulfovibrio species highly expressed genes for ethanol metabolism and extracellular electron transfer involving cytochromes, pili and flagella. Methanobacterium species in the aggregates also expressed genes for enzymes involved in reducing carbon dioxide to methane. Pure cultures demonstrated that the isolated Desulfovibrio sp. strain JY contributed to aggregate conductivity and directly transferred electrons to Methanothrix harundinacea, which is unable to use H2 or formate. Most importantly, further coculture studies indicated that Methanobacterium strain YSL might directly accept electrons from the Desulfovibrio strain JY for the reduction of carbon dioxide to methane in the aggregate. This finding suggested that the possibility of DIET by Desulfovibrio similar to Geobacter species in conductive methanogenic aggregates can not be excluded.


Assuntos
Desulfovibrio , Geobacter , Desulfovibrio/genética , Transporte de Elétrons , Elétrons , Geobacter/genética , Metano , Methanobacterium/genética
4.
Bioresour Technol ; 340: 125717, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34426232

RESUMO

A microbial fuel cell-photocatalysis system with a novel photocatalytic air-cathode (MFC-PhotoCat) was proposed for synergistic degradation of 2,4,6-trichlorophenol (TCP) with simultaneous electricity generation. Stable electricity generation of 350 mV was achieved during 130 days of operation. Besides, 50 mg L-1 TCP was completely degraded within 72 h, and the rate constant of 0.050 h-1 was 1.8-fold higher than MFC with air-cathode without N-TiO2 photocatalyst. Degradation pathway was proposed based on the intermediates detected and density functional theory (DFT) calculation, with two open-chain intermediates (2-chloro-4-keto-2-hexenedioic acid and hexanoic acid) detected. Furthermore, hierarchical cluster and PCoA revealed significant shifts of microbial community structures, with enriched exoelectrogen (55.2% of Geobacter) and TCP-degrading microbe (7.1% of Thauera) on the cathode biofilm as well as 61.8% of Pseudomonas in the culture solution. This study provides a promising strategy for synergic degradation of recalcitrant contaminants by intimate-coupling of MFC and photocatalysis.


Assuntos
Fontes de Energia Bioelétrica , Poluentes Ambientais , Geobacter , Eletricidade , Eletrodos
5.
Int J Mol Sci ; 22(16)2021 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-34445739

RESUMO

Environmental changes trigger the continuous adaptation of bacteria to ensure their survival. This is possible through a variety of signal transduction pathways involving chemoreceptors known as methyl-accepting chemotaxis proteins (MCP) that allow the microorganisms to redirect their mobility towards favorable environments. MCP are two-component regulatory (or signal transduction) systems (TCS) formed by a sensor and a response regulator domain. These domains synchronize transient protein phosphorylation and dephosphorylation events to convert the stimuli into an appropriate cellular response. In this review, the variability of TCS domains and the most common signaling mechanisms are highlighted. This is followed by the description of the overall cellular topology, classification and mechanisms of MCP. Finally, the structural and functional properties of a new family of MCP found in Geobacter sulfurreducens are revisited. This bacterium has a diverse repertoire of chemosensory systems, which represents a striking example of a survival mechanism in challenging environments. Two G. sulfurreducens MCP-GSU0582 and GSU0935-are members of a new family of chemotaxis sensor proteins containing a periplasmic PAS-like sensor domain with a c-type heme. Interestingly, the cellular location of this domain opens new routes to the understanding of the redox potential sensing signaling transduction pathways.


Assuntos
Quimiotaxia/fisiologia , Geobacter/fisiologia , Proteínas Quimiotáticas Aceptoras de Metil/fisiologia , Transdução de Sinais
6.
Environ Sci Technol ; 55(15): 10821-10831, 2021 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-34288663

RESUMO

Microbial reduction of Fe(III) minerals is a prominent process in redoximorphic soils and is strongly affected by organic matter (OM). We herein determined the rate and extent of microbial reduction of ferrihydrite (Fh) with either adsorbed or coprecipitated OM by Geobacter sulfurreducens. We focused on OM-mediated effects on electron uptake and alterations in Fh crystallinity. The OM was obtained from anoxic soil columns (effluent OM, efOM) and included-unlike water-extractable OM-compounds released by microbial activity under anoxic conditions. We found that organic molecules in efOM had generally no or only very low electron-accepting capacity and were incorporated into the Fh aggregates when coprecipitated with Fh. Compared to OM-free Fh, adsorption of efOM to Fh decelerated the microbial Fe(III) reduction by passivating the Fh surface toward electron uptake. In contrast, coprecipitation of Fh with efOM accelerated the microbial reduction, likely because efOM disrupted the Fh structure, as noted by Mössbauer spectroscopy. Additionally, the adsorbed and coprecipitated efOM resulted in a more sustained Fe(III) reduction, potentially because efOM could have effectively scavenged biogenic Fe(II) and prevented the passivation of the Fh surface by the adsorbed Fe(II). Fe(III)-OM coprecipitates forming at anoxic-oxic interfaces are thus likely readily reducible by Fe(III)-reducing bacteria in redoximorphic soils.


Assuntos
Compostos Férricos , Solo , Geobacter , Ferro , Minerais , Oxirredução
7.
Nat Commun ; 12(1): 4119, 2021 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-34226558

RESUMO

Northern peatlands are experiencing more frequent and severe fire events as a result of changing climate conditions. Recent studies show that such a fire-regime change imposes a direct climate-warming impact by emitting large amounts of carbon into the atmosphere. However, the fires also convert parts of the burnt biomass into pyrogenic carbon. Here, we show a potential climate-cooling impact induced by fire-derived pyrogenic carbon in laboratory incubations. We found that the accumulation of pyrogenic carbon reduced post-fire methane production from warm (32 °C) incubated peatland soils by 13-24%. The redox-cycling, capacitive, and conductive electron transfer mechanisms in pyrogenic carbon functioned as an electron snorkel, which facilitated extracellular electron transfer and stimulated soil alternative microbial respiration to suppress methane production. Our results highlight an important, but overlooked, function of pyrogenic carbon in neutralizing forest fire emissions and call for its consideration in the global carbon budget estimation.


Assuntos
Carbono/metabolismo , Elétrons , Metano/biossíntese , Incêndios Florestais , Bactérias , Biomassa , Dióxido de Carbono , Clima , Mudança Climática , Ecossistema , Incêndios , Geobacter , Laboratórios , Solo
8.
Environ Sci Technol ; 55(14): 10142-10151, 2021 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-34196176

RESUMO

Direct interspecies electron transfer (DIET) between microbial species prevails in some key microbial consortia. However, the electron transfer mechanism(s) in these consortia is controversial due to lack of efficient characterization methods. Here, we provide an in situ anaerobic spectroelectrochemical coculture cell (in situ ASCC) to induce the formation of DIET coculture biofilm on the interdigitated microelectrode arrays and characterize the electron transfer directly. Two typical Geobacter DIET cocultures, Geobacter metallireducens and wild-type Geobacter sulfurreducens (G.m&G.s) and G. metallireducens and a G. sulfurreducens strain deficient in citrate synthase (G.m&G.s-ΔgltA), were selected. In situ Raman and electrochemical Fourier transform infrared (FTIR) spectroscopy indicated that cytochromes are abundant in the electric syntrophic coculture. Cyclic voltammetry and potential step experiment revealed a diffusion-controlled electron transfer process and the electrochemical gating measurements further demonstrated a cytochrome-mediated electron transfer in the DIET coculture. Furthermore, the G.m&G.s-ΔgltA coculture displayed a higher redox conductivity than the G.m&G.s coculture, consistent with the existence of an intimate and efficient electrical connection between these two species. Our findings provide the first report of a redox-gradient-driven electron transport facilitated by c-type cytochromes in DIET coculture, supporting the model that DIET is mediated by cytochromes and suggest a platform to explore the other DIET consortia.


Assuntos
Geobacter , Técnicas de Cocultura , Citocromos/metabolismo , Transporte de Elétrons , Geobacter/metabolismo , Oxirredução
9.
J Phys Chem B ; 125(30): 8305-8312, 2021 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-34292748

RESUMO

Long-range electron transport has been widely and experimentally reported in Geobacter sulfurreducens pilus protein. However, a better understanding of the still undefined molecular arrangement can bring to light the role of key residues in this phenomenon. We propose a theoretical investigation of the electronic structure of aromatic residue groups in the protein through a classical molecular dynamics (MD) simulation, followed by a quantum mechanics/molecular mechanics (QM/MM) electronic study of different frames sampled from MD trajectories, an electrostatic potential and electron density analysis, an analysis of the density of states, and an investigation of hole formation through Dyson orbital calculations. We observe a highest occupied molecular orbital-lowest unoccupied molecular orbital (HOMO-LUMO) energy gap in the ranges of 1.4-2.3 eV and 2.9-3.3 eV and a less intense dipole moment along the aromatic residues in the presence of water in comparison to the system in vacuum. HOMO and LUMO electron densities highlight the occupation of one tyrosine residue in every representation for HOMO and a delocalization along two to three rings for LUMO. The results show how the electronic structure of the aromatic residues is sensitive to the ring arrangement and the surrounding environment. In our study, we observe that slight rearrangements in the fiber geometry can create temporary conditions for hole transfer.


Assuntos
Geobacter , Simulação de Dinâmica Molecular , Eletrônica , Fímbrias Bacterianas
10.
Chemosphere ; 283: 130983, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34153910

RESUMO

Biochar was proved as an electron shuttle to facilitate extracellular electron transfer (EET) of electrochemically active bacteria (EAB); however, its underlying mechanism was not fully understood. In this study, we aimed to further explore how the regulation of surface functional groups of biochar would affect the microbial iron reduction process of Geobacter sulfurreducens as a typical EAB. Two modified biochars were achieved after HNO3 (NBC) and NaBH4 (RBC) pretreatments, and a control biochar was produced after deionized water (WBC) washing. Results showed that WBC and RBC significantly accelerated microbial iron reduction of G. sulfurreducens PCA, while had no effect in the final Fe (II) minerals (e.g., vivianite and green rust (CO32-)). Besides, Brunauer-Emmett-Teller (BET) surface area, electron spin resonance (ESR) and electrochemical measurements showed that larger surface area, lower redox potential, and more redox-active groups (e.g., aromatic structures and quinone/hydroquinone moieties) in RBC explained its better electron transfer performance comparing to WBC. Interestingly, NBC completely suppressed the Fe (III) reduction process, mainly due to the production of reactive oxygen species which inhibited the growth of G. sulfurreducens PCA. Overall, this work paves a feasible way to regulate the surface functional groups for biochar, and comprehensively revealed its effect on EET process of microorganisms.


Assuntos
Geobacter , Carvão Vegetal , Transporte de Elétrons , Compostos Férricos , Ferro , Oxirredução
11.
Environ Sci Technol ; 55(12): 8258-8266, 2021 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-34096274

RESUMO

Anaerobic digestion (AD) and microbial electrochemical technologies (MET) can be combined in manifold ways. Recent studies show negative influences of AD effluents on the performance of pre-grown Geobacter spp.-dominated biofilm anodes. In this study, it was investigated how such biofilm anodes are affected by AD effluents. Therefore, experiments using AD effluents in different concentrations (0-100%) in combination with biofilms of different ages were performed. Furthermore, the activity of methanogens was inhibited and minimized by application of 2-bromoethanesulfonate (2-BES) and microfiltration, respectively. Biofilms pre-grown for 5 weeks show higher resistance against AD effluents compared to biofilms pre-grown for only 3 weeks. Nevertheless, adaptation of biofilms to AD effluents was not successful. Biofilm activity in terms of coulombic efficiency and maximum current density (jmax) dropped by factor 32.2 ± 3.2 and 38.9 ± 8.4, respectively. The application of 2-BES and microfiltration had positive effects on the biofilm activity. The results support the assumption that methanogens or further compounds not studied here, for example, protozoans, which may have been inhibited or removed by 2-BES application or microfiltration, have an immediate influence on the stability of Geobacter spp.-dominated biofilms and may limit their practical application in AD environments.


Assuntos
Fontes de Energia Bioelétrica , Geobacter , Anaerobiose , Biofilmes , Eletrodos
12.
BMC Genomics ; 22(1): 475, 2021 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-34171987

RESUMO

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.


Assuntos
Geobacter , Desulfuromonas , Eletrodos , Transporte de Elétrons , Compostos Férricos , Oxirredução
13.
Appl Environ Microbiol ; 87(17): e0070621, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34190605

RESUMO

A strain of Geobacter sulfurreducens, an organism capable of respiring solid extracellular substrates, lacking four of five outer membrane cytochrome complexes (extABCD+ strain) grows faster and produces greater current density than the wild type grown under identical conditions. To understand cellular and biofilm modifications in the extABCD+ strain responsible for this increased performance, biofilms grown using electrodes as terminal electron acceptors were sectioned and imaged using electron microscopy to determine changes in thickness and cell density, while parallel biofilms incubated in the presence of nitrogen and carbon isotopes were analyzed using NanoSIMS (nanoscale secondary ion mass spectrometry) to quantify and localize anabolic activity. Long-distance electron transfer parameters were measured for wild-type and extABCD+ biofilms spanning 5-µm gaps. Our results reveal that extABCD+ biofilms achieved higher current densities through the additive effects of denser cell packing close to the electrode (based on electron microscopy), combined with higher metabolic rates per cell compared to the wild type (based on increased rates of 15N incorporation). We also observed an increased rate of electron transfer through extABCD+ versus wild-type biofilms, suggesting that denser biofilms resulting from the deletion of unnecessary multiheme cytochromes streamline electron transfer to electrodes. The combination of imaging, physiological, and electrochemical data confirms that engineered electrogenic bacteria are capable of producing more current per cell and, in combination with higher biofilm density and electron diffusion rates, can produce a higher final current density than the wild type. IMPORTANCE Current-producing biofilms in microbial electrochemical systems could potentially sustain technologies ranging from wastewater treatment to bioproduction of electricity if the maximum current produced could be increased and current production start-up times after inoculation could be reduced. Enhancing the current output of microbial electrochemical systems has been mostly approached by engineering physical components of reactors and electrodes. Here, we show that biofilms formed by a Geobacter sulfurreducens strain producing ∼1.4× higher current than the wild type results from a combination of denser cell packing and higher anabolic activity, enabled by an increased rate of electron diffusion through the biofilms. Our results confirm that it is possible to engineer electrode-specific G. sulfurreducens strains with both faster growth on electrodes and streamlined electron transfer pathways for enhanced current production.


Assuntos
Biofilmes , Espaço Extracelular/metabolismo , Geobacter/química , Geobacter/fisiologia , Eletricidade , Eletrodos , Transporte de Elétrons , Espaço Extracelular/química , Geobacter/crescimento & desenvolvimento
14.
Biosensors (Basel) ; 11(6)2021 May 26.
Artigo em Inglês | MEDLINE | ID: mdl-34073192

RESUMO

Sustainable technologies for energy production and storage are currently in great demand. Bioelectrochemical systems (BESs) offer promising solutions for both. Several attempts have been made to improve carbon felt electrode characteristics with various pretreatments in order to enhance performance. This study was motivated by gaps in current knowledge of the impact of pretreatments on the enrichment and microbial composition of bioelectrochemical systems. Therefore, electrodes were treated with poly(neutral red), chitosan, or isopropanol in a first step and then fixed in microbial electrolysis cells (MECs). Four MECs consisting of organic substance-degrading bioanodes and methane-producing biocathodes were set up and operated in batch mode by controlling the bioanode at 400 mV vs. Ag/AgCl (3M NaCl). After 1 month of operation, Enterococcus species were dominant microorganisms attached to all bioanodes and independent of electrode pretreatment. However, electrode pretreatments led to a decrease in microbial diversity and the enrichment of specific electroactive genera, according to the type of modification used. The MEC containing isopropanol-treated electrodes achieved the highest performance due to presence of both Enterococcus and Geobacter. The obtained results might help to select suitable electrode pretreatments and support growth conditions for desired electroactive microorganisms, whereby performance of BESs and related applications, such as BES-based biosensors, could be enhanced.


Assuntos
Fontes de Energia Bioelétrica , Biofilmes , Fibra de Carbono , Eletrodos , Carbono , Eletrólise , Geobacter
15.
Water Res ; 200: 117270, 2021 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-34077836

RESUMO

The accumulation of volatile fatty acids, particularly propionic acid, significantly inhibits the efficiency of the anaerobic digestion system. In propionate degradation metabolism, the unfavorable thermodynamics of syntrophic reactions, strict ecological niche of syntrophic priopionate oxidizing bacteria, and slow metabolic rate of methanogens are regarded as major limitations. In this study, Geobacter sulfurreducens was co-cultured with Syntrophobacter fumaroxidans in bioelelectrochemical cells to analyze the propionate degradation process, impact factor, mechanism metabolic pathways, and electron transfer comprehensively. The results revealed that the syntroph S. fumaroxidans and syntrophic partner G. sulfurreducens achieved more efficient propionate degradation than the control group, comprising S. fumaroxidans and methanogens. Moreover, the carbon resource concentration and pH were both significantly correlated with propionate degradation (P < 0.01). The results further confirmed that G. sulfurreducen strengthened the consumption of H2 and acetate via direct interspecific electron transfer in propionate degradation. These findings indicate that G. sulfurreducens plays an unidentified functional role in propionate degradation.


Assuntos
Geobacter , Propionatos , Anaerobiose , Deltaproteobacteria , Metano
16.
ISME J ; 15(10): 3084-3093, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-33972726

RESUMO

Microbial corrosion of iron-based materials is a substantial economic problem. A mechanistic understanding is required to develop mitigation strategies, but previous mechanistic studies have been limited to investigations with relatively pure Fe(0), which is not a common structural material. We report here that the mechanism for microbial corrosion of stainless steel, the metal of choice for many actual applications, can be significantly different from that for Fe(0). Although H2 is often an intermediary electron carrier between the metal and microbes during Fe(0) corrosion, we found that H2 is not abiotically produced from stainless steel, making this corrosion mechanism unlikely. Geobacter sulfurreducens and Geobacter metallireducens, electrotrophs that are known to directly accept electrons from other microbes or electrodes, extracted electrons from stainless steel via direct iron-to-microbe electron transfer. Genetic modification to prevent H2 consumption did not negatively impact on stainless steel corrosion. Corrosion was inhibited when genes for outer-surface cytochromes that are key electrical contacts were deleted. These results indicate that a common model of microbial Fe(0) corrosion by hydrogenase-positive microbes, in which H2 serves as an intermediary electron carrier between the metal surface and the microbe, may not apply to the microbial corrosion of stainless steel. However, direct iron-to-microbe electron transfer is a feasible route for stainless steel corrosion.


Assuntos
Geobacter , Corrosão , Elétrons , Geobacter/genética , Ferro , Aço Inoxidável
17.
J Biol Chem ; 296: 100711, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33915126

RESUMO

Geobacter bacteria are able to transfer electrons to the exterior of the cell and reduce extracellular electron acceptors including toxic/radioactive metals and electrode surfaces, with potential applications in bioremediation or electricity harvesting. The triheme c-type cytochrome PpcA from Geobacter metallireducens plays a crucial role in bridging the electron transfer from the inner to the outer membrane, ensuring an effective extracellular electron transfer. This cytochrome shares 80% identity with PpcA from Geobacter sulfurreducens, but their redox properties are markedly different, thus determining the distinctive working redox potential ranges in the two bacteria. PpcA from G. metallireducens possesses two extra aromatic amino acids (Phe-6 and Trp-45) in its hydrophobic heme core, whereas PpcA from G. sulfurreducens has a leucine and a methionine in the equivalent positions. Given the different nature of these residues in the two cytochromes, we have hypothesized that the extra aromatic amino acids could be partially responsible for the observed functional differences. In this work, we have replaced Phe-6 and Trp-45 residues by their nonaromatic counterparts in PpcA from G. sulfurreducens. Using redox titrations followed by UV-visible and NMR spectroscopy we observed that residue Trp-45 shifted the redox potential range 33% toward that of PpcA from G. sulfurreducens, whereas Phe-6 produced a negligible effect. For the first time, it is shown that the inclusion of an aromatic residue at the heme core can modulate the working redox range in abundant periplasmic proteins, paving the way to engineer bacterial strains for optimal microbial bioelectrochemical applications.


Assuntos
Citocromos/química , Citocromos/metabolismo , Geobacter/citologia , Geobacter/enzimologia , Heme , Periplasma/enzimologia , Interações Hidrofóbicas e Hidrofílicas , Cinética , Modelos Moleculares , Oxirredução , Domínios Proteicos
18.
Appl Environ Microbiol ; 87(12): e0261720, 2021 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-33837010

RESUMO

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. IMPORTANCE Microbial fuel cells are a promising technology for energy generation from various waste types. However, the molecular mechanisms of microbial extracellular electron transfer to the electrode need to be elucidated. G. sulfurreducens is a common key player in electricity generation in mixed-culture microbial fuel cell systems and a model microorganism for the study of extracellular electron transfer. Outer membrane c-type cytochrome OmcZ is essential for an optimal current production by G. sulfurreducens. OmcZ proteolytic cleavage occurs during maturation, but the underlying mechanism is unknown. This study identifies a subtilisin-like protease, OzpA, which plays a role in cleaving OmcZ and generating the mature form of OmcZ (OmcZS). OzpA is essential for current production and, thus, the proteolytic maturation of OmcZ. This is a novel regulation of the c-type cytochrome for G. sulfurreducens extracellular electron transfer. This study also provides new insights into the design strategy and development of microbial extracellular electron transfer for an efficient energy conversion from chemical energy to electricity.


Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Fontes de Energia Bioelétrica , Geobacter/metabolismo , Serina Proteases/metabolismo , Eletricidade , Geobacter/genética , Mutação , Proteólise , Serina Proteases/genética
19.
Environ Sci Technol ; 55(9): 5806-5814, 2021 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-33835790

RESUMO

The nitroheterocyclic 3-nitro-1,2,4-triazol-5-one (NTO) is an ingredient of insensitive explosives increasingly used by the military, becoming an emergent environmental pollutant. Cometabolic biotransformation of NTO occurs in mixed microbial cultures in soils and sludges with excess electron-donating substrates. Herein, we present the unusual energy-yielding metabolic process of NTO respiration, in which the NTO reduction to 3-amino-1,2,4-triazol-5-one (ATO) is linked to the anoxic acetate oxidation to CO2 by a culture enriched from municipal anaerobic digester sludge. Cell growth was observed simultaneously with NTO reduction, whereas the culture was unable to grow in the presence of acetate only. Extremely low concentrations (0.06 mg L-1) of the uncoupler carbonyl cyanide m-chlorophenyl hydrazone inhibited NTO reduction, indicating that the process was linked to respiration. The ultimate evidence of NTO respiration was adenosine triphosphate production due to simultaneous exposure to NTO and acetate. Metagenome sequencing revealed that the main microorganisms (and relative abundances) were Geobacter anodireducens (89.3%) and Thauera sp. (5.5%). This study is the first description of a nitroheterocyclic compound being reduced by anaerobic respiration, shedding light on creative microbial processes that enable bacteria to make a living reducing NTO.


Assuntos
Bactérias , Nitrocompostos , Bactérias/genética , Geobacter , Respiração , Triazóis
20.
Appl Environ Microbiol ; 87(11)2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33771781

RESUMO

Over the past century, microbiologists have studied organisms in pure culture, yet it is becoming increasingly apparent that the majority of biological processes rely on multispecies cooperation and interaction. While little is known about how such interactions permit cooperation, even less is known about how cooperation arises. To study the emergence of cooperation in the laboratory, we constructed both a commensal community and an obligate mutualism using the previously noninteracting bacteria Shewanella oneidensis and Geobacter sulfurreducens Incorporation of a glycerol utilization plasmid (pGUT2) enabled S. oneidensis to metabolize glycerol and produce acetate as a carbon source for G. sulfurreducens, establishing a cross-feeding, commensal coculture. In the commensal coculture, both species coupled oxidative metabolism to the respiration of fumarate as the terminal electron acceptor. Deletion of the gene encoding fumarate reductase in the S. oneidensis/pGUT2 strain shifted the coculture with G. sulfurreducens to an obligate mutualism where neither species could grow in the absence of the other. A shift in metabolic strategy from glycerol catabolism to malate metabolism was associated with obligate coculture growth. Further targeted deletions in malate uptake and acetate generation pathways in S. oneidensis significantly inhibited coculture growth with G. sulfurreducens The engineered coculture between S. oneidensis and G. sulfurreducens provides a model laboratory system to study the emergence of cooperation in bacterial communities, and the shift in metabolic strategy observed in the obligate coculture highlights the importance of genetic change in shaping microbial interactions in the environment.IMPORTANCE Microbes seldom live alone in the environment, yet this scenario is approximated in the vast majority of pure-culture laboratory experiments. Here, we develop an anaerobic coculture system to begin understanding microbial physiology in a more complex setting but also to determine how anaerobic microbial communities can form. Using synthetic biology, we generated a coculture system where the facultative anaerobe Shewanella oneidensis consumes glycerol and provides acetate to the strict anaerobe Geobacter sulfurreducens In the commensal system, growth of G. sulfurreducens is dependent on the presence of S. oneidensis To generate an obligate coculture, where each organism requires the other, we eliminated the ability of S. oneidensis to respire fumarate. An unexpected shift in metabolic strategy from glycerol catabolism to malate metabolism was observed in the obligate coculture. Our work highlights how metabolic landscapes can be expanded in multispecies communities and provides a system to evaluate the evolution of cooperation under anaerobic conditions.


Assuntos
Geobacter/fisiologia , Interações Microbianas , Shewanella/fisiologia , Simbiose , Anaerobiose , Técnicas de Cocultura , Biologia Sintética
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