Proton translocation coupled to methanogenesis from methanol + hydrogen in Methanosarcina barkeri

Abstract

Addition of methanol to resting cells of Methanosarcina barkeri incubated under an atmosphere of molecular hydrogen resulted in an acidification of the medium. This acidification was not observed when H 2 was replaced by N 2 or air, or when the uncoupler tetrachlorosalicylanilide was present. 2-Bromoethanesulfonate completely inhibited both methanogenesis and proton extrusion. N,N'-Dicyclohexylcarbodiimide, an inhibitor of the proton-translocating ATPase in M. barkeri, did not affect proton extrusion. Therefore, it could be concluded that proton translocation was coupled to the terminal methylcoenzyme M methylreductase reaction and that it was not due to an H÷-translocating ATPase. A maximum value of 4 H + translocated per CH 4 formed was calculated.Methanogenesis; Proton translocation; (M. barkeri)

Analysis of the vhoGAC and vhtGAC Operons from Methanosarcina mazei Strain Gö1, Both Encoding a Membrane‐bound Hydrogenase and a Cytochrome b

“…In the case of M s mazei strain GO1, the liberation of protons in the periplasm and the consumption of protons during -O,S(CH,),SSHpoThrP reduction in the cytoplasm may contribute to the formation of a proton gradient. However, the production of scalar protons cannot be the only mechanism of energizing the cytoplasmic membrane because 34 H' are transferred/l heterodisulfide reduced in organisms belonging to the family Methanosarcinaceae (Blaut et al, 1987). In future studies, we will attempt to analyze the composition of the entire electron-transport chain with the aim to identify other possible electron carriers and the mechanism of the proton-translocating processes.…”

Immunocytochemical localization of the coenzyme F420-reducing hydrogenase in Methanosarcina barkeri Fusaro

“…DISCUSSION The energy metabolism of methanogenic bacteria has probably been most extensively studied in methanol-grown cells of M. barkeri Fusaro, primarily during methane formation from a substrate mixture of methanol and hydrogen, and all experimental data published to date indicate a chemiosmotic mechanism for energy conservation involving an electrochemical proton gradient (19). Moreover, it has been concluded that the electron transfer from H2 to CH3-S-CoM drives the translocation of protons across the cytoplasmic membrane (11), since the acidification of the medium by resting-cell suspensions of M. barkeri could not be detected in the presence of 2-bromoethanesulfonic acid, a potent inhibitor of the methylreductase reaction (reaction 1), or when H2 was replaced by N2 or air. This interpretation is strongly supported by data obtained in labeling experiments under similar experimental conditions (10), for under an atmosphere of H2 whole cells of M. barkeri oxidized only 1% of the added [14C]methanol to '4CO2 but reduced 80% to 14CH4, precluding a significant contribution to the generation of the proton electrochemical gradient by methanol oxidation.…”

“…This interpretation is strongly supported by data obtained in labeling experiments under similar experimental conditions (10), for under an atmosphere of H2 whole cells of M. barkeri oxidized only 1% of the added [14C]methanol to '4CO2 but reduced 80% to 14CH4, precluding a significant contribution to the generation of the proton electrochemical gradient by methanol oxidation. Since the electrons for the reduction of CH3-S-CoM during methane formation from methanol and H2 apparently originate from the oxidation of hydrogen (10,11), it was extremely intriguing to determine the cytological localization of the F420-hydrogenase in whole cells of M. barkeri. This enzyme has been shown to constitute 1.6% of the total cellular protein in methanol-grown cells, suggesting an important role in the metabolism of this methanogenic bacterium (17).…”

“…During methanogenesis from methanol and H2 by restingcell suspensions of methanol-grown Methanosarcina barkeri, ATP synthesis is stringently coupled to an electrochemical proton gradient and a functional DCCD (N,N'-dicyclohexylcarbodiimide)-sensitive ATP synthase, indicating a chemiosmotic mechanism for energy conservation (8,10,11). The central intermediate of methane formation from methanol is 2-(methylthio)ethanesulfonic acid (CH3-S-coenzyme M [CoM]) (19,39,43) which is reductively demrethylated with H2 to yield CH4 and 2-mercaptoethanesulfonic acid (CoM-SH) by the multicomponent methyl-coenzyme M methylreductase system (36,37).…”

Chemiosmotic energy conversion and the membrane ATPase of Methanolobus tindarius

“…As reported by Blaut and Gottschalk [2], methanogens can use the energy from substrate reduction for ATP synthesis by an electron-transport-driven mechanism. Moreover, proton extrusion coupled to methanogenesis has been demonstrated [3]. The methylreductase system has been discussed as a candidate for a membrane-associated proton pump which catalyses the exergonic reduction of a coenzymebound methyl group to methane [4, 51.…”