Showing papers on "Methanosarcina barkeri published in 1985"
TL;DR: In this article, the authors compared the metabolism of methanol and acetate when Methanosarcina barkeri was grown in the presence and absence of Desulfovibrio vulgaris and showed that sulfate-dependent interspecies H2 transfer from M. barkeri to D. vulgaris resulted in less methane production, increased CO2 formation, and sulfide formation from substrates not directly utilized by the sulfate reducer as electron donors for energy metabolism and growth.
Abstract: We compared the metabolism of methanol and acetate when Methanosarcina barkeri was grown in the presence and absence of Desulfovibrio vulgaris. The sulfate reducer was not able to utilize methanol or acetate as the electron donor for energy metabolism in pure culture, but was able to grow in coculture. Pure cultures of M. barkeri produced up to 10 μmol of H2 per liter in the culture headspace during growth on acetate or methanol. In coculture with D. vulgaris, the gaseous H2 concentration was ≤2 μmol/liter. The fractions of 14CO2 produced from [14C]methanol and 2-[14C]acetate increased from 0.26 and 0.16, respectively, in pure culture to 0.59 and 0.33, respectively, in coculture. Under these conditions, approximately 42% of the available electron equivalents derived from methanol or acetate were transferred and were utilized by D. vulgaris to reduce approximately 33 μmol of sulfate per 100 μmol of substrate consumed. As a direct consequence, methane formation in cocultures was two-thirds that observed in pure cultures. The addition of 5.0 mM sodium molybdate or exogenous H2 decreased the effects of D. vulgaris on the metabolism of M. barkeri. An analysis of growth and carbon and electron flow patterns demonstrated that sulfate-dependent interspecies H2 transfer from M. barkeri to D. vulgaris resulted in less methane production, increased CO2 formation, and sulfide formation from substrates not directly utilized by the sulfate reducer as electron donors for energy metabolism and growth.
168 citations
TL;DR: The present work has provided the first proof of a chemiosmotic mechanism of ATP synthesis in methanogenic bacteria using methanol and H 2 as a raw material for methanogenesis.
81 citations
TL;DR: Data support the hypothesis that in M. barkeri, Na+ is involved not in the process of ATP synthesis but in the first step of methanol oxidation, and concludes that cell extracts of M. barkingeri are not able to oxidize methanl.
Abstract: Methanogenesis from methanol by cell suspensions of Methanosarcina barkeri was inhibited by the uncoupler tetrachlorosalicylanilide. This inhibition was reversed by the addition of formaldehyde. 14C labeling experiments revealed that methanol served exclusively as the electron acceptor, whereas formaldehyde was mainly oxidized to CO2 under these conditions. These data support the hypothesis (M. Blaut and G. Gottschalk, Eur. J. Biochem. 141: 217-222, 1984) that the first step in methanol oxidation depends on the proton motive force or a product thereof. Cell extracts of M. barkeri converted methanol and formaldehyde to methane under an H2 atmosphere. Under an N2 atmosphere, however, formaldehyde was disproportionated to CH4 and CO2, whereas methanol was metabolized to a very small extent only, irrespective of the presence of ATP. It was concluded that cell extracts of M. barkeri are not able to oxidize methanol. In further experiments, the sodium dependence of methanogenesis and ATP formation by whole cells was investigated. Methane formation from methanol alone and the corresponding increase in the intracellular ATP content were strictly dependent on Na+. If, in contrast, methanol was utilized together with H2, methane and ATP were synthesized in the absence of Na+. The same is true for the disproportionation of formaldehyde to methane and carbon dioxide. From these experiments, it is concluded that in M. barkeri, Na+ is involved not in the process of ATP synthesis but in the first step of methanol oxidation.
64 citations
TL;DR: It was found that the sludge supernatant could be replaced with yeast extract, 6 mM bicarbonate-30% CO(2), and trace metals, and the antimetabolite sulfanilamide (20 mM) inhibited growth of and methanogenesis by Methanosarcina sp.
Abstract: Methanosarcina sp. strain TM-1, an acetotrophic, thermophilic methanogen isolated from an anaerobic sludge digestor, was originally reported to require an anaerobic sludge supernatant for growth. It was found that the sludge supernatant could be replaced with yeast extract (1 g/liter), 6 mM bicarbonate-30% CO2, and trace metals, with a doubling time on methanol of 14 h. For growth on either methanol or acetate, yeast extract could be replaced with CaCl2 · 2H2O (13.6 μM minimum) and the vitamin p-aminobenzoic acid (PABA, ca. 3 nM minimum), with a doubling time on methanol of 8 to 9 h. Filter-sterilized folic acid at 0.3 μM could not replace PABA. The antimetabolite sulfanilamide (20 mM) inhibited growth of and methanogenesis by Methanosarcina sp. strain TM-1, and this inhibition was reversed by the addition of 0.3 μM PABA. When a defined medium buffered with 20 mM N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid was used, it was shown that Methanosarcina sp. strain TM-1 required 6 mM bicarbonate-30% CO2 for optimal growth and methanogenesis from methanol. Cells growing on acetate were less dependent on bicarbonate-CO2. When we used a defined medium in which the only organic compounds present were methanol or acetate, nitrilotriacetic acid (0.2 mM), and PABA, it was possible to limit batch cultures of Methanosarcina sp. strain TM-1 for nitrogen at NH4+ concentrations at or below 2.0 mM, in marked contrast with Methanosarcina barkeri 227, which fixes dinitrogen when grown under NH4+ limitation.
57 citations
TL;DR: The pathway of acetate catabolism in Methanosarcina barkeri strain MS was studied by using a recently developed assay for methanogenesis from acetate by soluble enzymes in cell extracts to discuss the acetate uptake and rate-limiting transformation mechanisms in methane formation.
Abstract: The pathway of acetate catabolism in Methanosarcina barkeri strain MS was studied by using a recently developed assay for methanogenesis from acetate by soluble enzymes in cell extracts. Extracts incubated with [2-14C]acetate, hydrogen, and ATP formed 14CH4 and [14C]methyl coenzyme M as products. The apparent Km for acetate conversion to methane was 5 mM. In the presence of excess acetate, both the rate and duration of methane production was dependent on ATP. Acetyl phosphate replaced the cell extract methanogenic requirement for both acetate and ATP (the Km for ATP was 2 mM). Low concentrations of bromoethanesulfonic acid and cyanide, inhibitors of methylreductase and carbon monoxide dehydrogenase, respectively, greatly reduced the rate of methanogenesis. Precipitation of CO dehydrogenase in cell extracts by antibodies raised to 95% purified enzyme inhibited both CO dehydrogenase and acetate-to-methane conversion activity. The data are consistent with a model of acetate catabolism in which methylreductase, methyl coenzyme M, CO dehydrogenase, and acetate-activating enzymes are components. These results are discussed in relation to acetate uptake and rate-limiting transformation mechanisms in methane formation.
57 citations
TL;DR: The metabolic properties of Methanothrix concilii are closely related to those of Methanosarcina barkeri, and the origin of carbon atoms in glutamate, proline, and arginine showed that an incomplete tricarboxylic acid cycle operating in the oxidative direction was used for their biosynthesis.
Abstract: Biosynthetic pathways in Methanothrix concilii, a recently isolated aceticlastic methanogen, were studied by 13C-nuclear magnetic resonance spectroscopy. Labeling patterns of amino acids, lipids, and carbohydrates were determined. Similar to other methanogens, acetate was carboxylated to pyruvate, which was further converted to amino acids by various biosynthetic pathways. The origin of carbon atoms in glutamate, proline, and arginine clearly showed that an incomplete tricarboxylic acid cycle operating in the oxidative direction was used for their biosynthesis. Isoleucine was synthesized via citramalate, which is a typical route for methanogens. As with Methanosarcina barkeri, an extensive exchange of the label between the carboxyl group of acetate and CO2 was observed. Lipids predominantly contained diphytanyl chains, the labeling of which indicated that biosynthesis proceeded through mevalonic acid. Labeling of the C-1,6 of glucose from [2-13C]acetate is consistent with a glucogenic route for carbohydrate biosynthesis. Except for the different origins of the methyl group of methionine, the metabolic properties of Methanothrix concilii are closely related to those of Methanosarcina barkeri.
57 citations
TL;DR: In this paper, the capacity of Methanosarcina barkeri to grow diazotrophically was proved with a pure culture in mineral media with methanol, and the cell yields with N2 or NH4+ ions as nitrogen source were 2.2 g and 6.1 g dry weight, respectively.
Abstract: Methanosarcina barkeri cells were observed in ammonia-free anaerobic acetate enrichments for sulfate-reducing bacteria. The capacity of Methanosarcina to grow diazotrophically was proved with a pure culture in mineral media with methanol. The cell yields with N2 or NH4+ ions as nitrogen source were 2.2 g and 6.1 g dry weight, respectively, per mol of methanol. Growth experiments with 15N2 revealed that 84% of the cell nitrogen was derived from N2. Acetylene was highly toxic to Methanosarcina and only reduced at concentrations lower than 100 μmol dissolved per 1 of medium. Assimilation of N2 and reduction of acetylene were inhibited by NH4+ ions. The experiments show that N2 fixation occurs not only in eubacteria but also in archaebacteria. The ecological significance of diazotrophic growth of Methanosarcina is discussed.
52 citations
TL;DR: Results suggest that inhibition by NaCN was a result of its action as an inhibitor of in vivo CO dehydrogenase, and implicate CO dehydrogensase in the oxidation of acetate but not methanol methyl groups to CO2.
Abstract: NaCN was tested for its inhibitory effects on growth of and metabolism by Methanosarcina barkeri 227. NaCN (10 microM) inhibited catabolism of acetate methyl groups to CH4 and CO2 but did not inhibit methanogenesis from methanol, CO2, methylamine, or trimethylamine. NaCN also inhibited the assimilation of methanol or CO2 (as the sole carbon source) into cell carbon and stimulated the assimilation of acetate. These results suggest that inhibition by NaCN was a result of its action as an inhibitor of in vivo CO dehydrogenase. The results also implicate CO dehydrogenase in the oxidation of acetate but not methanol methyl groups to CO2.
48 citations
TL;DR: In this paper, a corrinoid enzyme was found to have a role in methyl group transfer to coenzyme M after C-C cleavage of acetate in Methanosarcina barkeri.
Abstract: Methane formation from acetate in cell suspensions of Methanosarcina barkeri was inhibited by low concentrations (5 μM) of propyl iodide. Inhibition was abolished by short exposure of the suspension to light which strongly indicates that a corrinoid enzyme is involved in methanogenesis from acetate. Propyl iodide (5μM) had no effect on the exchange reaction between the carboxyl group of acetate and 14CO2, and on methane formation from methanol, from H2 and methanol, or from H2 and CO2. These findings indicate that the proposed corrinoid enzyme has a role in methyl group transfer to coenzyme M after C-C cleavage of acetate.
46 citations
TL;DR: Production of methane by Methanosarcina barkeri from H(2)-CO(2) was studied in fed-batch culture under phosphate-limiting conditions, indicating the reversibility of phosphate depletion.
Abstract: Production of methane by Methanosarcina barkeri from H2-CO2 was studied in fed-batch culture under phosphate-limiting conditions. A transition in the kinetics of methanogenesis from an exponentially increasing rate to a constant rate was due to depletion of phosphate from the medium. The period of exponentially increasing rate of methanogenesis was extended by increasing the initial concentration of phosphate in the medium. Addition of phosphate during the constant period changed the kinetics to an exponentially increasing rate of methanogenesis, indicating the reversibility of phosphate depletion. The relation between methanogenesis and growth of M. barkeri was investigated by measuring the incorporation of phosphorus, supplied as KH232PO4, in the medium. At a low (1 μM) initial concentration of phosphate in the medium and during the constant period of methanogenesis, there was no net cell growth. At a higher (10 μM) initial concentration of phosphate, cell growth proceeded linearly with time after phosphate had been removed from the medium by uptake into cells.
36 citations
TL;DR: The immunological approach demonstrates that aminoacyl-tRNA synthetases do not belong to the group of enzymes in gene expression with high structural conservation and achieve the necessary fidelity by post-transfer proofreading, a corrective hydrolytic event after transfer to tRNAPhe.
TL;DR: Archaebacteria Methanogen Chromosomal protein is a major component of E. coli infection and its role in wound healing and wound healing is still under investigation.
Journal Article•
TL;DR: Methanosarcina barkeri strain FR-19 lysed spontaneously in substrate-depleted cultures and the addition of 0.3 M-sucrose prevented complete lysis and resulted in the formation of osmotically sensitive UV-fluorescent spheres.
Abstract: Methanosarcina barkeri strain FR-19 lysed spontaneously in substrate-depleted cultures. The addition of 0.3 M-sucrose prevented complete lysis and resulted in the formation of osmotically sensitive UV-fluorescent spheres. Electron microscopic examination showed that complete degradation of the cell wall occurred before the release of true protoplasts, which were stabilized by sucrose and glucose (0.2-0.3 M) but not by glycerol. Exponentially increasing methane production and regeneration of protoplasts could not be demonstrated.
TL;DR: In this system, 9 g/l of formate could be obtained from CO gas after 48 h of incubation at 37°C, pH 8.0, and Hypophosphite inhibited formate formation from CO plus H2O by about 80%.
Abstract: A preliminary study of formate production from CO plus H2O using the intact cells ofMethanosarcina barkeri was conducted. Formate production from CO gas required the participation of three enzymes, CO dehydrogenase, hydrogenase and formate dehydrogenase. Hypophosphite inhibited formate formation from CO plus H2O by about 80%. In this system, 9 g/l of formate could be obtained from CO gas after 48 h of incubation at 37°C, pH 8.0.
TL;DR: The Methanospirillum pattern closely resembles the patterns displayed by Methanobacterium species and is sensitive to the peptidyl-transferase inhibitors anisomycin and narciclasine, while the Methanosarcina barkeri ribosomes, on the other hand, are not affected by these 80S ribosome targeted inhibitors.
01 Jan 1985
TL;DR: The divergent cofactor composition of CO dehydrogenases as well as significant differences in the affinity for CO reflect the quite different functions that are fulfilled by these enzymes in the bacterial groupings mentioned.
Abstract: Carbon monoxide is oxidized by bacteria from different physiological groupings, including both, aerobes and anaerobes. The groups of CO-oxidizing bacteria are carboxydotrophs, methanotrophs, dinitrogen-fixers, acetogens, methanogens, phototrophs and sulfate- reducers. Some can use CO as sole carbon and energy source for growth (utilitarian CO-oxidizers) whereas the gas is only co-oxidized by others (non-utilitarian CO-oxidizers). Enzymes that Catalyze the oxidation of CO to CO2 may be subdivided according to whether they contain molybdenum or nickel as an integral component of their active center. To date molybdenum-containing CO dehydrogenases have been identified in Pseudomonas carboxydovorans, Pseudomonas carboxydohydrogena, Pseudomonas carboxydoflava and Bacillus schlegelii. In addition, these enzymes contain the molybdopterin of the molybdenum cofactor, flavin and two different iron-sulfur centers of the (2Fe-2S) type. Nickel-containing CO dehydrogenases have been found in Clostridium thermoaceticum, Acetobacterium woodii, Methanosarcina barkeri, and Desulfovibrio desulfuricans; most of them contain (4Fe-4S) centers, no flavin, and they are extremely sensitive to oxygen. The cofactor composition of CO dehydrogenases from aerobic bacteria appears to be much more complex than that of most enzymes from anaerobes. It is obvious, that CO dehydrogenases of aerobes are molybdenum iron-sulfur flavoproteins (molybdenum hydroxylases), whereas those of anaerobes are nickel iron-sulfur enzymes. The divergent cofactor composition of CO dehydrogenases as well as significant differences in the affinity for CO reflect the quite different functions that are fulfilled by these enzymes in the bacterial groupings mentioned.
TL;DR: Strain FAr9 is either an acetate permeability mutant or is defective in an activation step required for the catabolism and anabolism of acetate.
Abstract: Mutants of Methanosarcina barkeri 227 resistant to monofluoroacetate were isolated from monofluoroacetate-treated cultures. Mutant strain FAr9 was 100 times more resistant to monofluoroacetate than the wild-type strain and was deficient in carbon uptake and CH4 and CO2 production from methyl-labeled acetate. Methanol was assimilated at increased levels. Strain FAr9 was unable to shift from using methanol to using acetate for growth and exhibited increased sensitivity to growth inhibition by NaCN in methanol-containing complex medium. Unlike parent strain 227, acetate addition to methanol-containing media did not prevent NaCN inhibition. The specific activities of enzymes of exogenous acetate assimilation, CO dehydrogenase, and enzymes of the tricarboxylic acid cycle were similar for mutant and parent strain cell extracts. Mutation to monofluoroacetate resistance did not confer simultaneous resistance to 2-bromoethanesulfonate or pyruvate or alter propionate uptake. We conclude that strain FAr9 is either an acetate permeability mutant or is defective in an activation step required for the catabolism and anabolism of acetate.
TL;DR: Chemical and immunochemical analyses of hybridomas constructed with spleen cells from mice immunized against Methanosarcina barkeri 227 showed that antigen 227 is a high-molecular-weight structure composed mainly of one carbohydrate, glucose, and small amounts of amino acids, which suggests that this molecule is associated with a lipid moiety.
Abstract: Hybridomas were constructed with spleen cells from mice immunized against Methanosarcina barkeri 227. The reaction with the resulting monoclonal antibodies identified two antigenic determinants. Determinant 8A is present in M. barkeri 227, where it is accessible to antibody on whole bacterial cells. 8A is undetectable in (or absent from) M. barkeri R1M3, an immunologically closely related strain. Determinant 8C is present in both strains, but with M. barkeri 227 it is found only in extracts and cannot be demonstrated in whole cells. It therefore appears to be hidden. A soluble form of antigen 8A (antigen 227) was obtained treating whole M. barkeri 227 cells with absolute methanol. This antigen was further purified by affinity chromatography with antibody 8A. Chemical and immunochemical analyses of these preparations showed that antigen 227 is a high-molecular-weight (4 X 10(5)) structure composed mainly of one carbohydrate, glucose, and small amounts of amino acids. Its solubility properties suggest that this molecule is associated with a lipid moiety.
TL;DR: The Desulfobulbus sp. with Methanosarcina barkeri 227 was able to produce CH4 from propionate in the presence of sulfate, if a sufficient amount of ferrous iron was added to the media in order to trap the soluble sulfides produced from sulfate as mentioned in this paper.
Abstract: The association of Desulfobulbus sp. with Methanosarcina barkeri 227 was able to produce CH4 from propionate in the presence of sulfate, if a sufficient amount of ferrous iron was added to the media in order to trap the soluble sulfides produced from sulfate. In the absence of ferrous iron, soluble sulfides inhibited the acetoclastic reaction. Attempts to cultivate Desulfobulbus sp. with H2-utilising methanogenic bacteria in the absence of sulfate did not succeed.
01 Jan 1985
TL;DR: The association of Desulfobulbus sp.
Abstract: 1. SUMMARY The association of Desulfobulbus sp. with Methanosarcina barkeri 227 was able to produce CH 4 from propionate in the presence of sulfate, if a sufficient amount of ferrous iron was added to the media in order to trap the soluble sulfides produced from-stllfate. In the absence of ferrous iront soluble sulfides inhibited the acetoclastic re action. Attempts to cultivate Desulfobulbus sp. with H 2 -utilising methanogenic bacteria in the absence of sulfate did noLsucceed.