scholarly journals Direct Electron Transfer between the frhAGB-Encoded Hydrogenase and Thioredoxin Reductase in the Nonmethanogenic Archaeon Thermococcus onnurineus NA1

2020 ◽  
Vol 86 (6) ◽  
Author(s):  
Hae-Chang Jung ◽  
Jae Kyu Lim ◽  
Tae-Jun Yang ◽  
Sung Gyun Kang ◽  
Hyun Sook Lee
Keyword(s):  
Electron Transfer ◽  
Electron Donor ◽  
Direct Electron Transfer ◽  
Thioredoxin Reductase ◽  
Thermococcus Onnurineus Na1 ◽  
Content Type ◽  
Protein Target ◽  
Redox Partner ◽  
Significant Step

ABSTRACT To date, NAD(P)H, ferredoxin, and coenzyme F420 have been identified as electron donors for thioredoxin reductase (TrxR). In this study, we present a novel electron source for TrxR. In the hyperthermophilic archaeon Thermococcus onnurineus NA1, the frhAGB-encoded hydrogenase, a homolog of the F420-reducing hydrogenase of methanogens, was demonstrated to interact with TrxR in coimmunoprecipitation experiments and in vitro pulldown assays. Electrons derived from H2 oxidation by the frhAGB-encoded hydrogenase were transferred to TrxR and reduced Pdo, a redox partner of TrxR. Interaction and electron transfer were observed between TrxR and the heterodimeric hydrogenase complex (FrhAG) as well as the heterotrimeric complex (FrhAGB). Hydrogen-dependent reduction of TrxR was 7-fold less efficient than when NADPH was the electron donor. This study not only presents a different type of electron donor for TrxR but also reveals new functionality of the frhAGB-encoded hydrogenase utilizing a protein as an electron acceptor. IMPORTANCE This study has importance in that TrxR can use H2 as an electron donor with the aid of the frhAGB-encoded hydrogenase as well as NAD(P)H in T. onnurineus NA1. Further studies are needed to explore the physiological significance of this protein. This study also has importance as a significant step toward understanding the functionality of the frhAGB-encoded hydrogenase in a nonmethanogen; the hydrogenase can transfer electrons derived from oxidation of H2 to a protein target by direct contact without the involvement of an electron carrier, which is distinct from the mechanism of its homologs, F420-reducing hydrogenases of methanogens.

scholarly journals Iron Corrosion via Direct Metal-Microbe Electron Transfer

mBio ◽  
2019 ◽  
Vol 10 (3) ◽  
Author(s):  
Hai-Yan Tang ◽  
Dawn E. Holmes ◽  
Toshiyuki Ueki ◽  
Paola A. Palacios ◽  
Derek R. Lovley
Keyword(s):  
Electron Transfer ◽  
Electron Donor ◽  
Environmental Concern ◽  
Direct Electron Transfer ◽  
Anaerobic Respiration ◽  
Electrical Contacts ◽  
Geobacter Sulfurreducens ◽  
Donor Strain ◽  
Iron Corrosion ◽  
Content Type

ABSTRACTThe concept that anaerobic microorganisms can directly accept electrons from Fe(0) has been controversial because direct metal-microbe electron transfer has previously only been indirectly inferred. Fe(0) oxidation was studied withGeobacter sulfurreducensstrain ACL, an autotrophic strain that was previously shown to grow with electrons derived from a graphite cathode as the sole electron donor. Strain ACL grew with Fe(0) as the sole electron donor and fumarate as the electron acceptor. However, it appeared that at least a portion of the electron transfer was via H2produced nonenzymatically from the oxidation of Fe(0) to Fe(II). H2, which accumulated in abiotic controls, was consumed during the growth of strain ACL, the cells were predominately planktonic, and genes for the uptake hydrogenase were highly expressed. Strain ACLHFwas constructed to prevent growth on H2or formate by deleting the genes for the uptake of hydrogenase and formate dehydrogenases from strain ACL. Strain ACLHFalso grew with Fe(0) as the sole electron donor, but H2accumulated in the culture, and cells heavily colonized Fe(0) surfaces with no visible planktonic growth. Transcriptomics suggested that the outer surfacec-type cytochromes OmcS and OmcZ were important during growth of strain ACLHFon Fe(0). Strain ACLHFdid not grow on Fe(0) if the gene for either of these cytochromes was deleted. The specific attachment of strain ACLHFto Fe(0), coupled with requirements for known extracellular electrical contacts, suggest that direct metal-microbe electron transfer is the most likely option for Fe(0) serving as an electron donor.IMPORTANCEThe anaerobic corrosion of iron structures is expensive to repair and can be a safety and environmental concern. It has been known for over 100 years that the presence of anaerobic respiratory microorganisms can accelerate iron corrosion. Multiple studies have suggested that there are sulfate reducers, methanogens, and acetogens that can directly accept electrons from Fe(0) to support sulfate or carbon dioxide reduction. However, all of the strains studied can also use H2as an electron donor for growth, which is known to be abiotically produced from Fe(0). Furthermore, no proteins definitely shown to function as extracellular electrical contacts with Fe(0) were identified. The studies described here demonstrate that direct electron transfer from Fe(0) can support anaerobic respiration. They also map out a simple genetic approach to the study of iron corrosion mechanisms in other microorganisms. A better understanding of how microorganisms promote iron corrosion is expected to lead to the development of strategies that can help reduce adverse impacts from this process.

In Vitro Evaluation of Miniaturized Amperometric Enzyme Sensor Based on the Direct Electron Transfer Principle for Continuous Glucose Monitoring

2022 ◽  
pp. 193229682110706
Author(s):  
Yutaro Inoue ◽  
Yasuhide Kusaka ◽  
Kotaro Shinozaki ◽  
Inyoung Lee ◽  
Koji Sode
Keyword(s):  
Electron Transfer ◽  
Continuous Glucose Monitoring ◽  
Glucose Sensor ◽  
Direct Electron Transfer ◽  
Glucose Dehydrogenase ◽  
Glucose Monitoring ◽  
Reference Electrode ◽  
Enzyme Sensor

Background: The bacterial derived flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase (FADGDH) is the most promising enzyme for the third-generation principle-based enzyme sensor for continuous glucose monitoring (CGM). Due to the ability of the enzyme to transfer electrons directly to the electrode, recognized as direct electron transfer (DET)-type FADGDH, although no investigation has been reported about DET-type FADGDH employed on a miniaturized integrated electrode. Methods: The miniaturized integrated electrode was formed by sputtering gold (Au) onto a flexible film with 0.1 mm in thickness and divided into 3 parts. After an insulation layer was laminated, 3 openings for a working electrode, a counter electrode and a reference electrode were formed by dry etching. A reagent mix containing 1.2 × 10−4 Unit of DET-type FADGDH and carbon particles was deposited. The long-term stability of sensor was evaluated by continuous operation, and its performance was also evaluated in the presence of acetaminophen and the change in oxygen partial pressure (pO2) level. Results: The amperometric response of the sensor showed a linear response to glucose concentration up to 500 mg/dL without significant change of the response over an 11-day continuous measurement. Moreover, the effect of acetaminophen and pO2 on the response were negligible. Conclusions: These results indicate the superb potential of the DET-type FADGDH-based sensor with the combination of a miniaturized integrated electrode. Thus, the described miniaturized DET-type glucose sensor for CGM will be a promising tool for effective glycemic control. This will be further investigated using an in vivo study.

Bioconversion of Carbon Monoxide to Formate Using Artificially Designed Carbon Monoxide:Formate Oxidoreductase in Hyperthermophilic Archaea

2021 ◽  
Author(s):  
Jae Kyu Lim ◽  
Ji-In Yang ◽  
Yun Jae Kim ◽  
Yeong-Jun Park ◽  
Yong Hwan Kim
Keyword(s):  
Carbon Monoxide ◽  
Electron Transfer ◽  
Fusion Protein ◽  
Direct Electron Transfer ◽  
Co2 Reduction ◽  
Hyperthermophilic Archaea ◽  
Production Activity ◽  
Formate Production

Abstract Ferredoxin-dependent metabolic engineering of electron transfer circuits has been developed to enhance redox efficiency in the field of synthetic biology, e.g., for hydrogen production and for reduction of flavoproteins or NAD(P)+. Here, we present the bioconversion of carbon monoxide (CO) gas to formate via a synthetic CO:formate oxidoreductase (CFOR), designed as an enzyme complex for direct electron transfer between noninteracting CO dehydrogenase and formate dehydrogenase using an electron-transferring Fe-S fusion protein. The CFOR-introduced Thermococcus onnurineus mutant strains showed CO-dependent formate production in vivo and in vitro. The formate production rate from purified CFOR complex and specific formate productivity from the bioreactor were 348 ± 34 μmol/mg/min and 90.2 ± 20.4 mmol/g-cells/h, respectively. The CO-dependent CO2 reduction/formate production activity of synthetic CFOR was confirmed, indicating that direct electron transfer between two unrelated dehydrogenases was feasible via mediation of the FeS-FeS fusion protein.

Distinct electron transfer from ferredoxin–thioredoxin reductase to multiple thioredoxin isoforms in chloroplasts

2017 ◽  
Vol 474 (8) ◽  
pp. 1347-1360 ◽  
Author(s):  
Keisuke Yoshida ◽  
Toru Hisabori
Keyword(s):  
Electron Transfer ◽  
Redox Regulation ◽  
Thioredoxin Reductase ◽  
Reducing Power ◽  
Redox Potentials ◽  
Regulation Network ◽  
Electron Transfers ◽  
Kinetics Of

Thiol-based redox regulation is considered to support light-responsive control of various chloroplast functions. The redox cascade via ferredoxin–thioredoxin reductase (FTR)/thioredoxin (Trx) has been recognized as a key to transmitting reducing power; however, Arabidopsis thaliana genome sequencing has revealed that as many as five Trx subtypes encoded by a total of 10 nuclear genes are targeted to chloroplasts. Because each Trx isoform seems to have a distinct target selectivity, the electron distribution from FTR to multiple Trxs is thought to be the critical branch point for determining the consequence of chloroplast redox regulation. In the present study, we aimed to comprehensively characterize the kinetics of electron transfer from FTR to 10 Trx isoforms. We prepared the recombinant FTR protein from Arabidopsis in the heterodimeric form containing the Fe–S cluster. By reconstituting the FTR/Trx system in vitro, we showed that FTR prepared here was enzymatically active and suitable for uncovering biochemical features of chloroplast redox regulation. A series of redox state determinations using the thiol-modifying reagent, 4-acetamido-4′-maleimidylstilbene-2,2′-disulfonate, indicated that all chloroplast Trx isoforms are commonly reduced by FTR; however, significantly different efficiencies were evident. These differences were apparently correlated with the distinct midpoint redox potentials among Trxs. Even when the experiments were performed under conditions of hypothetical in vivo stoichiometry of FTR and Trxs, a similar trend in distinguishable electron transfers was observed. These data highlight an aspect of highly organized circuits in the chloroplast redox regulation network.

scholarly journals Uniform and Pitting Corrosion of Carbon Steel byShewanella oneidensisMR-1 under Nitrate-Reducing Conditions

2018 ◽  
Vol 84 (12) ◽  
pp. e00790-18 ◽  
Author(s):  
Robert B. Miller ◽  
Kenton Lawson ◽  
Anwar Sadek ◽  
Chelsea N. Monty ◽  
John M. Senko
Keyword(s):  
Electron Transfer ◽  
Carbon Steel ◽  
Electron Donor ◽  
Pitting Corrosion ◽  
Nitrate Reduction ◽  
Steel Corrosion ◽  
Nitrite Accumulation ◽  
Content Type ◽  
Microbially Influenced Corrosion ◽  
Reducing Conditions

ABSTRACTDespite observations of steel corrosion in nitrate-reducing environments, processes of nitrate-dependent microbially influenced corrosion (MIC) remain poorly understood and difficult to identify. We evaluated carbon steel corrosion byShewanella oneidensisMR-1 under nitrate-reducing conditions using a split-chamber/zero-resistance ammetry (ZRA) technique. This approach entails the deployment of two metal (carbon steel 1018 in this case) electrodes into separate chambers of an electrochemical split-chamber unit, where the microbiology or chemistry of the chambers can be manipulated. This approach mimics the conditions of heterogeneous metal coverage that can lead to uniform and pitting corrosion. The current between working electrode 1 (WE1) and WE2 can be used to determine rates, mechanisms, and, we now show, extents of corrosion. WhenS. oneidensiswas incubated in the WE1 chamber with lactate under nitrate-reducing conditions, nitrite transiently accumulated, and electron transfer from WE2 to WE1 occurred as long as nitrite was present. Nitrite in the WE1 chamber (withoutS. oneidensis) induced electron transfer in the same direction, indicating that nitrite cathodically protected WE1 and accelerated the corrosion of WE2. WhenS. oneidensiswas incubated in the WE1 chamber without an electron donor, nitrate reduction proceeded, and electron transfer from WE2 to WE1 also occurred, indicating that the microorganism could use the carbon steel electrode as an electron donor for nitrate reduction. Our results indicate that under nitrate-reducing conditions, uniform and pitting carbon steel corrosion can occur due to nitrite accumulation and the use of steel-Fe(0) as an electron donor, but conditions of sustained nitrite accumulation can lead to more-aggressive corrosive conditions.IMPORTANCEMicrobially influenced corrosion (MIC) causes damage to metals and metal alloys that is estimated to cost over $100 million/year in the United States for prevention, mitigation, and repair. While MIC occurs in a variety of settings and by a variety of organisms, the mechanisms by which microorganisms cause this damage remain unclear. Steel pipe and equipment may be exposed to nitrate, especially in oil and gas production, where this compound is used for corrosion and “souring” control. In this paper, we show uniform and pitting MIC under nitrate-reducing conditions and that a major mechanism by which it occurs is via the heterogeneous cathodic protection of metal surfaces by nitrite as well as by the microbial oxidation of steel-Fe(0).

Concerted Ion and Electron Transfer Across Electronically Conductive Polymer Membranes

1992 ◽  
Vol 293 ◽  
Author(s):  
Charles R. Martin ◽  
Del R. Lawson ◽  
Wenbin Liang
Keyword(s):  
Electron Transfer ◽  
Electron Donor ◽  
Direct Electron Transfer ◽  
Conductive Polymer ◽  
Electron Transfer Reaction ◽  
Reduced Form ◽  
Free Standing ◽  
Transfer Processes ◽  
Donor Acceptor ◽  
Synthetic Metal

AbstractWe describe in this paper an experiment involving an electronically conductive polymer (ECP) which, to our knowledge, has not been described previously. A free-standing ECP (polypyrrole)-based membrane separates a solution of an electron donor from a solution of an electron acceptor. Because the ECP is both electronically and anionically conductive, the membrane can transport electrons from the donor solution to the acceptor solution, and anions in the opposite direction, such that a sustainable electron-transfer reaction is driven across the ECP membrane. We demonstrate such transmembrane electron/ion-transfer processes using both an inorganic and a biochemical electron donor/acceptor system. The biochemical case is of particular interest because we show that the reduced form of the enzyme glucose oxidase can give its electrons directly to the synthetic metal. Direct electron transfer is usually not possible at inorganic metals.

scholarly journals Antioxidant Activity of the Yeast Mitochondrial One-Cys Peroxiredoxin Is Dependent on Thioredoxin Reductase and Glutathione In Vivo

2009 ◽  
Vol 29 (11) ◽  
pp. 3229-3240 ◽  
Author(s):  
Darren Greetham ◽  
Chris M. Grant
Keyword(s):  
Oxidative Stress ◽  
Antioxidant Activity ◽  
Thioredoxin Reductase ◽  
Disulfide Bridge ◽  
Cysteine Residue ◽  
In Vitro Assays ◽  
Redox Partner ◽  
Thioredoxin System

ABSTRACT Peroxiredoxins are ubiquitous enzymes which protect cells against oxidative stress. The first step of catalysis is common to all peroxiredoxins and results in oxidation of a conserved peroxidatic cysteine residue to sulfenic acid. This forms an intermolecular disulfide bridge in the case of 2-Cys peroxiredoxins, which is a substrate for the thioredoxin system. 1-Cys Prx's contain a peroxidatic cysteine but do not contain a second conserved cysteine residue, and hence the identity of the in vivo reduction system has been unclear. Here, we show that the yeast mitochondrial 1-Cys Prx1 is reactivated by glutathionylation of the catalytic cysteine residue and subsequent reduction by thioredoxin reductase (Trr2) coupled with glutathione (GSH). This novel mechanism does not require the usual thioredoxin (Trx3) redox partner of Trr2 for antioxidant activity, although in vitro assays show that the Trr2/Trx3 and Trr2/GSH systems exhibit similar capacities for supporting Prx1 catalysis. Our data also indicate that mitochondria are a main target of cadmium-induced oxidative stress and that Prx1 is particularly required to protect against mitochondrial oxidation. This study demonstrates a physiological reaction mechanism for 1-Cys peroxiredoxins and reveals a new role in protection against mitochondrial heavy metal toxicity.

scholarly journals Flavin Electron Shuttles Dominate Extracellular Electron Transfer by Shewanella oneidensis

mBio ◽  
2013 ◽  
Vol 4 (1) ◽  
Author(s):  
Nicholas J. Kotloski ◽  
Jeffrey A. Gralnick
Keyword(s):  
Electron Transfer ◽  
Direct Electron Transfer ◽  
Shewanella Oneidensis ◽  
Phenotypic Characterization ◽  
Extracellular Electron Transfer ◽  
Wild Type ◽  
Content Type ◽  
Electron Shuttling ◽  
Electron Shuttles ◽  
Set Up

ABSTRACT Shewanella oneidensis strain MR-1 is widely studied for its ability to respire a diverse array of soluble and insoluble electron acceptors. The ability to breathe insoluble substrates is defined as extracellular electron transfer and can occur via direct contact or by electron shuttling in S. oneidensis. To determine the contribution of flavin electron shuttles in extracellular electron transfer, a transposon mutagenesis screen was performed with S. oneidensis to identify mutants unable to secrete flavins. A multidrug and toxin efflux transporter encoded by SO_0702 was identified and renamed bfe (bacterial flavin adenine dinucleotide [FAD] exporter) based on phenotypic characterization. Deletion of bfe resulted in a severe decrease in extracellular flavins, while overexpression of bfe increased the concentration of extracellular flavins. Strains lacking bfe had no defect in reduction of soluble Fe(III), but these strains were deficient in the rate of insoluble Fe(III) oxide reduction, which was alleviated by the addition of exogenous flavins. To test a different insoluble electron acceptor, graphite electrode bioreactors were set up to measure current produced by wild-type S. oneidensis and the Δbfe mutant. With the same concentration of supplemented flavins, the two strains produced similar amounts of current. However, when exogenous flavins were not supplemented to bioreactors, bfe mutant strains produced significantly less current than the wild type. We have demonstrated that flavin electron shuttling accounts for ~75% of extracellular electron transfer to insoluble substrates by S. oneidensis and have identified the first FAD transporter in bacteria. IMPORTANCE Extracellular electron transfer by microbes is critical for the geochemical cycling of metals, bioremediation, and biocatalysis using electrodes. A controversy in the field was addressed by demonstrating that flavin electron shuttling, not direct electron transfer or nanowires, is the primary mechanism of extracellular electron transfer employed by the bacterium Shewanella oneidensis. We have identified a flavin adenine dinucleotide transporter conserved in all sequenced Shewanella species that facilitates export of flavin electron shuttles in S. oneidensis. Analysis of a strain that is unable to secrete flavins demonstrated that electron shuttling accounts for ~75% of the insoluble extracellular electron transfer capacity in S. oneidensis.

scholarly journals Thioredoxin Dependent Changes in the Redox States of FurA from Anabaena sp. PCC 7120

Antioxidants ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 913
Author(s):  
Jorge Guío ◽  
María Teresa Bes ◽  
Mónica Balsera ◽  
Laura Calvo-Begueria ◽  
Emma Sevilla ◽  
...  
Keyword(s):  
Reductase Activity ◽  
Thioredoxin Reductase ◽  
Zinc Ion ◽  
Exchange Reactions ◽  
Redox Partner ◽  
Redox Partners ◽  
Anabaena Sp ◽  
Pcc 7120

FurA is a multifunctional regulator in cyanobacteria that contains five cysteines, four of them arranged into two CXXC motifs. Lack of a structural zinc ion enables FurA to develop disulfide reductase activity. In vivo, FurA displays several redox isoforms, and the oxidation state of its cysteines determines its activity as regulator and its ability to bind different metabolites. Because of the relationship between FurA and the control of genes involved in oxidative stress defense and photosynthetic metabolism, we sought to investigate the role of type m thioredoxin TrxA as a potential redox partner mediating dithiol-disulfide exchange reactions necessary to facilitate the interaction of FurA with its different ligands. Both in vitro cross-linking assays and in vivo two-hybrid studies confirmed the interaction between FurA and TrxA. Light to dark transitions resulted in reversible oxidation of a fraction of the regulator present in Anabaena sp. PCC7120. Reconstitution of an electron transport chain using E. coli NADPH-thioredoxin-reductase followed by alkylation of FurA reduced cysteines evidenced the ability of TrxA to reduce FurA. Furthermore, the use of site-directed mutants allowed us to propose a plausible mechanism for FurA reduction. These results point to TrxA as one of the redox partners that modulates FurA performance.

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