Identification of a vanadate-sensitive, membrane-bound ATPase in the archaebacterium Methanococcus voltae

Dharmavaram, Rita M.; Konisky, J

doi:10.1128/jb.169.9.3921-3925.1987

Cited by 29 publications

(11 citation statements)

References 42 publications

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“…While we previously reported the presence of a membrane-associated P-type ATPase in M. voltae, our studies failed to identify an ATPase with the properties characteristic of an ATP synthase (11)(12)(13). In the present study, we used an antibody raised against the subunit of the H+-translocating ATP synthase purified from the archaea S. acidocaldarius (29,30) Cells equilibrated with 24Na' as described in the legend to Fig.…”

Section: Resultscontrasting

confidence: 43%

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Energy transduction in the methanogen Methanococcus voltae is based on a sodium current

Dybas

Konisky

1992

J Bacteriol

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We provide experimental support for the proposal that ATP production in Methanococcus voltae, a methanogenic member of the archaea, is based on an energetic system in which sodium ions, not protons, are the coupling ions. We show that when grown at a pH of 6.0, 7.1, or 8.2, M. voltae cells maintain a membrane potential of approximately -150 mV. The cells maintain a transmembrane pH gradient (pH,. -pHo.t) of -0.1, -0.2, and -0.2, respectively, values not favorable to the inward movement of protons. The cells maintain a transmembrane sodium concentration gradient (sodiumo.,/sodium1.) of 1.2, 3.4, and 11.6, respectively. While the protonophore 3,3',4',5-tetrachlorosalicylanilide inhibits ATP formation in cells grown at pH 6.5, neither ATP formation nor growth is inhibited in cells grown in medium at pH 8.2. We show that when grown at pH 8.2, cells synthesize ATP in the absence of a favorably oriented proton motive force. Whether grown at pH 6.5 or pH 8.2, M. voltae extrudes Na+ via a primary pump whose activity does not depend on a proton motive force. The addition of protons to the cells leads to a harmaline-sensitive efflux of Na+ and vice versa, indicating the presence of Na+/H+ antiporter activity and, thus, a second mechanism for the translocation of Na+ across the cell membrane. M. voltae contains a membrane component that is immunologically related to the H+-translocating ATP synthase of the archaeabacterium Sulfolobus acidocaldanius. Since we demonstrated that ATP production can be driven by an artificially imposed membrane potential only in the presence of sodium ions, we propose that ATP production in M. voltae is mediated by an Na+-translocating ATP synthase whose function is coupled to a sodium motive force that is generated through a primary Na* pump.Evidence has been obtained with whole cells of Methanosarcina barkeri and with whole cells, protoplasts, and crude membrane vesicles of methanogenic bacterium strain Gol that reduction of the heterodisulfide of CH3-S-CoM [2-(methylthio)ethanesulfonate-methylcoenzyme M] and 7-mercaptoheptanoylthreoninephosphate leads to the generation of a proton motive force (PMF) that is coupled to the synthesis of ATP via an H+-translocating ATP synthase (3,4,9,10,23,32). In contrast, results obtained in studies directed towards elucidating the energetics of ATP formation in Methanococcus voltae have shown that methane formation and ATP synthesis can occur in the presence of the protonophore SF6847 and have led to the conclusion that the maintenance of a PMF is not required for ATP synthesis in this marine methanogen (7). On the basis of these findings, a novel and intriguing scheme of substrate-level phosphorylation was considered possible (27).To study the mechanism of ATP formation in M. voltae, we decided to reexamine the possibility that ATP formation was mediated by a chemiosmotically based mechanism by considering a mechanism in which the coupling ion is not a proton. Since growth, amino acid transport, and ATP production in whole cells of M. voltae had been shown...

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confidence: 43%

“…ATP levels in energized cells were determined by withdrawal of 500-,ul samples and extraction with a solution of 10% trichloroacetic acid-4 mM EDTA. Extracts were washed three times with 3 volumes of diethyl ether prior to the ATP level determination by the luciferin-luciferase assay as previously described (12).…”

Section: Methodsmentioning

confidence: 99%

Energy transduction in the methanogen Methanococcus voltae is based on a sodium current

Dybas

Konisky

1992

J Bacteriol

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“…6 shows the effects of azide and nitrate on the ATPase activity. Azide, a potent inhibitor of F-type ATPases, lowers the activity only [8] and other P-type ATPases, vanadate was found to decrease the activity of the Methanolobus ATPase only at concentrations three orders of magnitude higher.…”

Section: Characterisation Of the Membrane-bound Atpasementioning

confidence: 99%

“…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.ATPases as possible catalysts of ATP synthesis have been found in the membranes of several methanogens [6-101; one of these has been identified as being vanadate-sensitive [8].Ion-transducing membrane ATPases can be divided into three classes, the P (or El-E2) type of plasma membranes, the V type of vacuolar membranes and the F type of chloroplasts, mitochondria and bacterial membranes [I I]. Members of these classes differ in their physiological role and their sensitivity against ionic [12] and antibiotic [13] inhibitors.…”

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Chemiosmotic energy conversion and the membrane ATPase of Methanolobus tindarius

Scheel

Schäfer

1990

European Journal of Biochemistry

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Electron transport phosphorylation has been demonstrated to drive ATP synthesis for the methanogenic archaebacterium Methanolobus tindarius: Protonophores evoked uncoupler effects and lowered the membrane potential Ay. Under the influence of N,N'-dicyclohexylcarbodiimide [(cHxN),C] the membrane potential increased while methanol turnover was inhibited. 2-Bromoethanesulfonate, an inhibitor of methanogenesis, had no effect on the membrane potential but, like (CHXN)~C and protonophores, decreased the intracellular ATP concentration.Labeling experiments with ( C H X N )~'~C showed membranes to contain a proteolipid, with a molecular mass of 5.5 kDa, that resembles known (cHxN),C-binding proteins of Fo-F1 ATPases. The (cHxN)2-sensitive membrane ATPase hydrolysed Mg . ATP at a pH optimum of 5.0 with a K , (ATP) of 2.5 mM (V = 77 mujmg). It was inhibited competitively by ADP; Ki (ADP) = 0.65 mM. Azide or vanadate caused no significant loss in ATPase activity, but millimolar concentrations of nitrate showed an inhibitory effect, suggesting a relationship to ATPases from vacuolar membranes. In contrast, no inhibition occurred in the presence of bafilomycin Al.The ATPase was extractable with EDTA at low salt concentrations. The purified enzyme consists of four different subunits, CI (67 kDa), , h' (52 kDa), y (20 kDa) and 6 ( < 10 kDa), as determined from SDS gel electrophoresis.Among archaebacteria the methanogens represent a diverse group of strictly anaerobic organisms producing methane from CO,, acetate, formate, methanol and methylamines by disproportionation or by reduction with molecular hydrogen (for a review see [l]). 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.ATPases as possible catalysts of ATP synthesis have been found in the membranes of several methanogens [6-101; one of these has been identified as being vanadate-sensitive [8].Ion-transducing membrane ATPases can be divided into three classes, the P (or El-E2) type of plasma membranes, the V type of vacuolar membranes and the F type of chloroplasts, mitochondria and bacterial membranes [I I]. Members of these classes differ in their physiological role and their sensitivity against ionic [12] and antibiotic [13] inhibitors.

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Structure and Mechanism of F ₀ F ₁ ‐Type ATP Synthases and ATPases

Penefsky

Cross

1991

Advances in Enzymology - And Related Areas of Molecular Biology

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Identification of a vanadate-sensitive, membrane-bound ATPase in the archaebacterium Methanococcus voltae

Cited by 29 publications

References 42 publications

Energy transduction in the methanogen Methanococcus voltae is based on a sodium current

Energy transduction in the methanogen Methanococcus voltae is based on a sodium current

Chemiosmotic energy conversion and the membrane ATPase of Methanolobus tindarius

Structure and Mechanism of F ₀ F ₁ ‐Type ATP Synthases and ATPases

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Identification of a vanadate-sensitive, membrane-bound ATPase in the archaebacterium Methanococcus voltae

Cited by 29 publications

References 42 publications

Energy transduction in the methanogen Methanococcus voltae is based on a sodium current

Energy transduction in the methanogen Methanococcus voltae is based on a sodium current

Chemiosmotic energy conversion and the membrane ATPase of Methanolobus tindarius

Structure and Mechanism of F 0 F 1 ‐Type ATP Synthases and ATPases

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Structure and Mechanism of F ₀ F ₁ ‐Type ATP Synthases and ATPases