ATP synthesis coupled to electron transfer from H<sub>2</sub> to the heterodisulfide of 2‐mercaptoethanesulfonate and 7‐mercaptoheptanoylthreonine phosphate in vesicle preparations of the methanogenic bacterium strain Gö1

Peinemann, S.; Hedderich, Reiner; Blaut, Michaël; Thauer, Rudolf K.; Gottschalk, Gerhard

doi:10.1016/0014-5793(90)80704-m

Cited by 34 publications

(20 citation statements)

References 13 publications

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“…Redox titrations were performed by the methods of Dutton (9) and Wilson (39) in a customdesigned cuvette (path length, 1 cm; Ochs, Bovenden, Germany) at 25°C. Redox potentials were measured with a micro-redox-electrode type Pt 5700 A (combination electrode; Schott, Hofheim, Germany) in the presence of the following redox mediators: 2,6-diaminodurol (Em7 = +260 mV), 33 ,uM; 1,2-naphthoquinone (Em 7 = +143 mV), 34 ,uM; phenazine methosulfate (Em7 = +80 mV), 18 ,uM; phenazine ethosulfate (Em 7 = +55 mV), 16 ,uM; duroquinone (Em,7 = +5 mV), 33 ,uM; pyocyanine (Emn7 = -34 mV), 4 R,M; 2,3-dimethyl-1,4-naphthoquinone (Em 7 = -80 mV), 83 ,uM; 2-hydroxy-1,4-naphthoquinone (Em;7 -152 mV), 31 ,uM; anthraquinone-2,6-disulfonate (Em 7 = -184 mV), 18 ,uM; anthraquinone-2-sulfonate (Em 7 = -224 mV), 18 ,uM; phenosafranine (Em 7 = -252 mV), 0.23 ,uM; acridinium chloride (Em,7 = -313 mV), 19 ,uM; diquat (Em7 = -349 mV), 5 ,uM; methylviologen (Em 7 = -449 mV), 0.4 ,uM. To obtain a fast response, the platinum electrode was activated by successive incubations in hydroquinone (in 40 mM sodium oxalate buffer, pH 4.0) and Na2SO3 (in H20) followed by sandpapering prior to each titration.…”

Section: Growthmentioning

confidence: 99%

“…The methanogenic pathways leading to the conversion of these substrates have been unraveled in the last two decades, and they involve a number of unique cofactors (37). The penultimate step of methanogenesis, the reduction of the heterodisulfide of coenzyme M and 7-mercaptoheptanoylthreonine phosphate (CoM-S-S-HTP) with either H2 or reduced coenzyme F420 (F420H2) as a reductant, was shown in Methanosarcina strain Gol to be coupled to electron transport phosphorylation (7,34 Membrane preparations. For the spectroscopic and potentiometric characterization of cytochromes, membranes were prepared under aerobic conditions.…”

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Characterization of cytochromes from Methanosarcina strain Göl and their involvement in electron transport during growth on methanol

Kamlage

Blaut

1992

J Bacteriol

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Methanosarcina strain Gol was tested for the presence of cytochromes. Low-temperature spectroscopy, hemochrome derivative spectroscopy, and redox titration revealed the presence of two b-type (b559 and b564) and two c-type (Cs47 and c552) cytochromes in membranes from Methanosarcina strain Gol. The midpoint potentials determined were Em 7 =-135 + 5 and -240 + 11 mV (b-type cytochromes) and Em 7 =-140 10 and -230 10 mV (c-type cytochromes). The protoheme IX and the heme c contents were 0.21 to 0.24 and 0.09 to 0.28 ,Imol/g of membrane protein, respectively. No cytochromes were detectable in the cytoplasmic fraction. Of various electron donors and acceptors tested, only the reduced form of coenzyme F420 (coenzyme F420H2) and the heterodisulfide of coenzyme M and 7-mercaptoheptanoylthreonine phosphate (CoM-S-S-HTP) were capable of reducing and oxidizing the cytochromes at a high rate, respectively. Addition of CoM-S-S-HTP to reduced cytochromes and subsequent low-temperature spectroscopy revealed the oxidation of cytochrome b564. On the basis of these results, we suggest that one or several cytochromes participate in the coenzyme F420H2-dependent reduction of the heterodisulfide.Methanogenic bacteria gain metabolic energy by coupling the conversion of a number of simple substrates such as H2-CO2, formate, methylamines, methanol, and acetate to the generation of ATP. The methanogenic pathways leading to the conversion of these substrates have been unraveled in the last two decades, and they involve a number of unique cofactors (37). The penultimate step of methanogenesis, the reduction of the heterodisulfide of coenzyme M and 7-mercaptoheptanoylthreonine phosphate (CoM-S-S-HTP) with either H2 or reduced coenzyme F420 (F420H2) as a reductant, was shown in Methanosarcina strain Gol to be coupled to electron transport phosphorylation (7,34 Membrane preparations. For the spectroscopic and potentiometric characterization of cytochromes, membranes were prepared under aerobic conditions. The freshly harvested cells were washed twice with 50 mM potassium phosphate buffer, pH 7.0, containing 0.2 M sucrose and 10 mM MgSO4. After suspension in 50 mM potassium phosphate buffer, pH 7.0 (KP buffer), and addition of a few crystals of DNasev the cells were lysed by passing them through an Aminco French pressure cell at 138 MPa. The resulting crude extract was centrifuged at 10,000 x g for 15 min to remove unbroken cells and cell debris. The supernatant was diluted to 150 ml with KP buffer and centrifuged at 120,000 x g for 2 h. The supernatant (cytoplasmic fraction) was carefully removed and discarded. The membrane pellet was resuspended in the same buffer and centrifuged at 10,000 x g for 15 min to pellet the residual sulfides apparently adhering to the membranes. The supernatant from this low-speed centrifugation was subsequently subjected to ultracentrifugation at 120,000 x g for 30 min. These steps (resuspension of the resulting pellet, low-speed centrifugation, and ultracentrifugation) were repeated two more times. The final m...

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Section: Growthmentioning

confidence: 99%

mentioning

confidence: 99%

Characterization of cytochromes from Methanosarcina strain Göl and their involvement in electron transport during growth on methanol

Kamlage

Blaut

1992

J Bacteriol

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“…CH3-S-CoM + HTP-SH CH4 + CoM-S-S-HTP [4] CoM-S-S-HTP + H2 CoM-SH + HTP-SH [5] Experimental evidence was provided that the reduction of the heterodisulfide (CoM-S-S-HTP) (reaction 5) might be the actual energy-conserving reaction (14).…”

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confidence: 99%

Reduced coenzyme F420: heterodisulfide oxidoreductase, a proton- translocating redox system in methanogenic bacteria.

Deppenmeier

Blaut

Mahlmann

et al. 1990

Proc. Natl. Acad. Sci. U.S.A.

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Washed everted vesicles of the methanogenic bacterium strain Gol were found to couple the F42OH2-dependent heterodisulfide reduction with the transfer of protons across the membrane into the lumen of the everted vesicles. The transmembrane electrochemical potential of protons thereby generated was shown to be competent in driving ATP synthesis from ADP + Pi, exhibiting a stoichiometry of 2 H' translocated or 0.4 ATP synthesized per F42AH2 oxidized.This enzyme system exhibits the phenomenon of coupling and uncoupling and represents a different kind of electron transport chain with the heterodisulflde of 2-mercaptoethanesulfonate and 7-mercaptoheptanoylthreonine phosphate as terminal electron acceptor. The heterodisulfide and methane are formed in the methyl coenzyme M reductase reaction. The reducing equivalents are derived from reduced coenzyme F420, which represents an analogue of NADH + H' in other respiratory chains. It is assumed that the proton-translocating oxidoreductase discovered in strain Gol is of principal importance to all methanogenic bacteria not utilizing H2.With the discovery of methanogens and the elucidation of their specialized, strictly anaerobic metabolism, the question was posed whether this group of organisms gains ATP by substrate-level phosphorylation, such as clostridia or streptococci, or whether the process of methanogenesis is somehow coupled with the generation of a transmembrane gradient of protons (1). The pathways involved in methane formation from the few substrates utilized [C02 + H2, formate, methanol, methylamines, and acetate (2)] have been worked out

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“…The reduction of CoM-S-S-COB with H, (Peinemann et al, 1990;Deppenmeier et al, 1991) and F,,,H, (Deppenmeier et al, 1990) has been shown to be coupled with energy conservation via the chemiosmotic mechanism (Deppenmeier et al, 1996).…”

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Heterodisulfide Reductase from Methanol‐Grown Cells of Methanosarcina Barkeri is not a Flavoenzyme

Künkel

Vaupel

Heim

et al. 1997

European Journal of Biochemistry

106

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Heterodisulfide reductase from methanol-grown cells of Methanosarcina barkeri (MbHdrDE) is a membrane-bound enzyme composed of a 46-kDa subunit MbHdrD and a 23-kDa subunit MbHdrE. The enzyme has been shown to contain 0.6 mol heme and 20 mol Fe/S per mol heterodimer. In addition, substoichiometric amounts of FAD, thought to be an essential component of the active enzyme, were detected. We have now obtained preparations of active heterodisulfide reductase in high yields completely devoid of a flavin. Cloning and sequencing of the genes encoding MbHdrD and MbHdrE, which were found to form a transcription unit hdrED, revealed that both subunits also lack an FAD-binding motif. MbHdr thus differs from heterodisulifde reductase from Methanobacterium thermoautotrophicum (MtHdr), which is a flavo iron-sulfur protein composed of the subunits MtHdrA (80 kDa), MtHdrB (36 kDa) and MtHdrC (21 kDa), the subunit HdrA harboring the flavin-binding site. Sequence comparisons revealed that the N-terminal third of MbHdrD, which contained two sequence motifs for [4Fe-4S] clusters, is similar to MtHdrC and that the C-terminal two thirds of MbHdrD are similar to MtHdrB. Thus, MbHdrD and MtHdrBC are structurally equivalent subunits. MbHdrE shows sequence similarity to b-type cytochromes, in agreement with the finding that this subunit contains a heme. These and other results indicate that MbHdrD harbors the active site of heterodisulfide reduction and that a flavin is not involved in catalysis. Since MbHdrD contains only iron-sulfur clusters, a mechanism of disulfide reduction involving one electron rather than two electron-transfer reactions has to be considered such as operative in ferredoxin :thioredoxin reductases from chloroplasts and cyanobacteria.

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ATP synthesis coupled to electron transfer from H₂ to the heterodisulfide of 2‐mercaptoethanesulfonate and 7‐mercaptoheptanoylthreonine phosphate in vesicle preparations of the methanogenic bacterium strain Gö1

Cited by 34 publications

References 13 publications

Characterization of cytochromes from Methanosarcina strain Göl and their involvement in electron transport during growth on methanol

Characterization of cytochromes from Methanosarcina strain Göl and their involvement in electron transport during growth on methanol

Reduced coenzyme F420: heterodisulfide oxidoreductase, a proton- translocating redox system in methanogenic bacteria.

Heterodisulfide Reductase from Methanol‐Grown Cells of Methanosarcina Barkeri is not a Flavoenzyme

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ATP synthesis coupled to electron transfer from H2 to the heterodisulfide of 2‐mercaptoethanesulfonate and 7‐mercaptoheptanoylthreonine phosphate in vesicle preparations of the methanogenic bacterium strain Gö1

Cited by 34 publications

References 13 publications

Characterization of cytochromes from Methanosarcina strain Göl and their involvement in electron transport during growth on methanol

Characterization of cytochromes from Methanosarcina strain Göl and their involvement in electron transport during growth on methanol

Reduced coenzyme F420: heterodisulfide oxidoreductase, a proton- translocating redox system in methanogenic bacteria.

Heterodisulfide Reductase from Methanol‐Grown Cells of Methanosarcina Barkeri is not a Flavoenzyme

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ATP synthesis coupled to electron transfer from H₂ to the heterodisulfide of 2‐mercaptoethanesulfonate and 7‐mercaptoheptanoylthreonine phosphate in vesicle preparations of the methanogenic bacterium strain Gö1