An Electron-bifurcating Caffeyl-CoA Reductase

Bertsch, Johannes; Parthasarathy, Anutthaman; Buckel, Wolfgañg; Müller, Volker

doi:10.1074/jbc.m112.444919

Cited by 78 publications

(105 citation statements)

References 51 publications

(53 reference statements)

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“…In dissimilatory sulfate reduction, a function in coupling of cytoplasmic redox reactions to membrane-associated redox reactions has been proposed (16,69). The HdrABC complex appears to be an electron-bifurcating module used over and over again in different contexts, as is the EtfAB complex (70), which is used in many anaerobes as an electronbifurcating module in crotonyl-CoA reduction (71), caffeyl-CoA reduction (72), and lactate oxidation (73). This hypothesis still remains to be experimentally verified.…”

Section: Discussionmentioning

confidence: 99%

Evidence for a Hexaheteromeric Methylenetetrahydrofolate Reductase in Moorella thermoacetica

et al. 2014

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c Moorella thermoacetica can grow with H 2 and CO 2 , forming acetic acid from 2 CO 2 via the Wood-Ljungdahl pathway. All enzymes involved in this pathway have been characterized to date, except for methylenetetrahydrofolate reductase (MetF). We report here that the M. thermoacetica gene that putatively encodes this enzyme, metF, is part of a transcription unit also containing the genes hdrCBA, mvhD, and metV. MetF copurified with the other five proteins encoded in the unit in a hexaheteromeric complex with an apparent molecular mass in the 320-kDa range. The 40-fold-enriched preparation contained per mg protein 3.1 nmol flavin adenine dinucleotide (FAD), 3.4 nmol flavin mononucleotide (FMN), and 110 nmol iron, almost as predicted from the primary structure of the six subunits. It catalyzed the reduction of methylenetetrahydrofolate with reduced benzyl viologen but not with NAD(P)H in either the absence or presence of oxidized ferredoxin. It also catalyzed the reversible reduction of benzyl viologen with NADH (diaphorase activity). Heterologous expression of the metF gene in Escherichia coli revealed that the subunit MetF contains one FMN rather than FAD. MetF exhibited 70-fold-higher methylenetetrahydrofolate reductase activity with benzyl viologen when produced together with MetV, which in part shows sequence similarity to MetF. Heterologously produced HdrA contained 2 FADs and had NAD-specific diaphorase activity. Our results suggested that the physiological electron donor for methylenetetrahydrofolate reduction in M. thermoacetica is NADH and that the exergonic reduction of methylenetetrahydrofolate with NADH is coupled via flavin-based electron bifurcation with the endergonic reduction of an electron acceptor, whose identity remains unknown.

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Section: Discussionmentioning

confidence: 99%

Evidence for a Hexaheteromeric Methylenetetrahydrofolate Reductase in Moorella thermoacetica

et al. 2014

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“…The major issue addressed in this work was whether or not the butyryl-CoA dehydrogenase from C. difficile belongs to the recently described novel subfamily of bifurcating enzymes (25,26,40) capable of coupling the exergonic reduction of crotonyl-CoA by NADH with the endergonic reduction of ferredoxin by NADH. In previous studies, ferredoxin produced as a product of crotonylCoA reduction was directly consumed by hydrogenase in order to release hydrogen, which was quantified for a description of enzyme activity (25).…”

Section: Discussionmentioning

confidence: 99%

Effect of an Oxygen-Tolerant Bifurcating Butyryl Coenzyme A Dehydrogenase/Electron-Transferring Flavoprotein Complex from Clostridium difficile on Butyrate Production in Escherichia coli

et al. 2013

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The butyrogenic genes from Clostridium difficile DSM 1296 T have been cloned and expressed in Escherichia coli. The enzymes acetyl-coenzyme A (CoA) C-acetyltransferase, 3-hydroxybutyryl-CoA dehydrogenase, crotonase, phosphate butyryltransferase, and butyrate kinase and the butyryl-CoA dehydrogenase complex composed of the dehydrogenase and two electron-transferring flavoprotein subunits were individually produced in E. coli and kinetically characterized in vitro. While most of these enzymes were measured using well-established test systems, novel methods to determine butyrate kinase and butyryl-CoA dehydrogenase activities with respect to physiological function were developed. Subsequently, the individual genes were combined to form a single plasmid-encoded operon in a plasmid vector, which was successfully used to confer butyrate-forming capability to the host. In vitro and in vivo studies demonstrated that C. difficile possesses a bifurcating butyryl-CoA dehydrogenase which catalyzes the NADH-dependent reduction of ferredoxin coupled to the reduction of crotonyl-CoA also by NADH. Since the reoxidation of ferredoxin by a membrane-bound ferredoxin:NAD ؉ -oxidoreductase enables electron transport phosphorylation, additional ATP is formed. The butyryl-CoA dehydrogenase from C. difficile is oxygen stable and apparently uses oxygen as a cooxidant of NADH in the presence of air. These properties suggest that this enzyme complex might be well suited to provide butyryl-CoA for solventogenesis in recombinant strains. The central role of bifurcating butyryl-CoA dehydrogenases and membrane-bound ferredoxin:NAD oxidoreductases (Rhodobacter nitrogen fixation [RNF]), which affect the energy yield of butyrate fermentation in the clostridial metabolism, is discussed. G enome sequencing of organisms provides information regarding the distribution of genes encoding biotechnologically important metabolic pathways. This is true for the clostridial butyrogenic pathway, which converts acetyl-conenzyme A (CoA)-the terminal oxidation product of glucose via glycolysis-to butyrate. Genes encoding enzymes from this pathway are widespread in genome-sequenced clostridia and related species (1-8). In spite of the central importance of butyrate-forming genes in these organisms, only individual enzymes from a comparably small selection of organisms have been purified and carefully studied in the past (9-15). In particular, Clostridium acetobutylicum enzymes were of great interest due to the organism's capability of producing acetone and butanol (16)(17)(18). Driven by the urgent need to replace oil-derived fossil fuels, genetic engineering of microbes for production of butan-1-ol, a promising alternative transportation fuel, is proceeding worldwide (16,(18)(19)(20)(21)(22)(23)(24).A long-known metabolic route such as the butyrate pathway (Fig. 1A) can suddenly gain fundamentally new meanings when hitherto-unknown properties of individual enzymes within this pathway become known. While for decades the role of butyrate formation was generally a...

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“…For example, it is thought that energy conservation during hydrogen-dependent caffeate respiration in A. woodii is governed by three FBEB complexes working in concert; the membrane associated Rnf, the soluble [FeFe]-hydrogenase, and the soluble caffeyl-CoA reductase-Etf complex responsible for oxidation of NADH and reduction of ferredoxin [38]. FBEB reactions metabolically 'hard-wire' the redox state of the different cofactor pools and electron carrier proteins.…”

Section: Energy Conservation In Acetogensmentioning

confidence: 99%

Trash to treasure: production of biofuels and commodity chemicals via syngas fermenting microorganisms

Latif¹,

Zeidan²,

Nielsen³

et al. 2014

Current Opinion in Biotechnology

177

125

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An Electron-bifurcating Caffeyl-CoA Reductase

Abstract: Background:Energy conservation by Na ϩ -translocating ferredoxin oxidation requires a low potential ferredoxin.

Cited by 78 publications

References 51 publications

Evidence for a Hexaheteromeric Methylenetetrahydrofolate Reductase in Moorella thermoacetica

Evidence for a Hexaheteromeric Methylenetetrahydrofolate Reductase in Moorella thermoacetica

Effect of an Oxygen-Tolerant Bifurcating Butyryl Coenzyme A Dehydrogenase/Electron-Transferring Flavoprotein Complex from Clostridium difficile on Butyrate Production in Escherichia coli

Trash to treasure: production of biofuels and commodity chemicals via syngas fermenting microorganisms

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