Methanosarcina barkeri

About: Methanosarcina barkeri is a research topic. Over the lifetime, 703 publications have been published within this topic receiving 32151 citations.

Characterization of Energy-Conserving Hydrogenase B in Methanococcus maripaludis

Abstract: Hydrogenotrophic methanococci specialize in utilizing H2 as an electron donor, and these organisms possess six different Ni-Fe hydrogenases. These enzymes include two F420--reducing hydrogenases, two non-F420-reducing hydrogenases, and two membrane-bound hydrogenases (Eha and Ehb [5]). The F420-reducing hydrogenases reduce coenzyme F420, which subsequently reduces methenyltetrahydromethanopterin and methylenetetrahydromethanopterin, intermediates in the pathway of methanogenesis. In Methanococcus voltae, the F420-reducing hydrogenase is also reported to reduce the 2-mercaptoethanesulfonate:7-mercaptoheptanoylthreonine phosphate heterodisulfide formed in the final step of methanogenesis (2). In contrast, Methanothermobacter marburgensis utilizes the non-F420-reducing hydrogenase to reduce the heterodisulfide (22, 25). The two membrane-bound hydrogenases couple the chemiosmotic energy of ion gradients to H2 oxidation and ferredoxin reduction. In the aceticlastic methanogen Methanosarcina barkeri, the homologous enzyme is called energy conserving hydrogenase or Ech and performs a variety of physiological functions, including the generation of a proton motive force during CO oxidation and concomitant proton reduction in aceticlastic methanogenesis and the generation of low potential electron donors for CO2 reduction to formylmethanofuran in the first step of methanogenesis and the reductive carboxylation of acetyl coenzyme A (acetyl-CoA) to pyruvate in carbon assimilation (11, 12). In the hydrogenotrophic methanogens, it is predicted that the two energy-conserving hydrogenases (Eha and Ehb) have distinct roles (26). The Ehb appears to reduce low potential electron carriers utilized in autotrophic CO2 fixation (16). Anabolic enzymes likely to be coupled to Ehb in this manner include (i) the carbon monoxide dehydrogenase/acetyl-CoA synthase (CODH/ACS) and the pyruvate oxidoreductase (Por), which catalyze the first two steps of carbon assimilation; (ii) the α-ketoglutarate oxidoreductase (Kor), which catalyzes the final step in the incomplete reductive tricarboxylic acid cycle; and (iii) the indolepyruvate oxidoreductase (Ior) and the 2-oxoisovalerate oxidoreductase (Vor), which are involved in amino acid biosynthesis from aryl and branched-chain acids, respectively. Support for these conclusions comes in large part from the phenotype of an M. maripaludis ehb gene replacement mutant S40, which was only capable of limited growth in the absence of acetate and amino acids (16). Furthermore, expression of CODH/ACS, Por, and Vor were significantly upregulated in the mutant, providing further evidence for a role of Ehb in these processes (16). In contrast, there is no direct evidence for the role of Eha. By analogy with the Methanosarcina Ech, it could be involved in generating reducing equivalents for the reduction of CO2 to formylmethanofuran. Alternatively, hydrogenotrophic methanogens may have an alternative method of CO2 reduction (27), and Eha could have another function entirely. In spite of some functional similarities between the Ech of the aceticlastic methanogens and Eha or Ehb of hydrogenotrophs, the structures of their operons are very different (Fig. ​(Fig.1).1). Based upon sequence comparisons, all of these membrane-bound hydrogenases possess conserved large and small hydrogenase subunits, a 2[4Fe-4S] ferredoxin, and an integral membrane ion translocator (3, 8, 26). Otherwise, the structures are very different. The purified Ech from Methanosarcina barkeri contains six polypeptides encoded by the six genes of the ech operon (8, 11). The Eha and Ehb hydrogenases have never been purified. The eha and ehb operons from the hydrogenotrophic methanogen Methanothermobacter thermautotrophicus comprise 20 and 17 genes, respectively (23, 26). Most of these genes are predicted to encode transmembrane proteins, although there are also several polyferredoxins and hydrophilic proteins (26). Many of these genes are not homologous to the M. barkeri ech genes. The Methanococcus maripaludis genome contains homologs to the M. thermautotrophicus eha and ehb genes, although only nine of the ehb genes are contiguous on the genome (Fig. ​(Fig.1).1). In the present study, the Ehb from the hydrogenotrophic methanogen Methanococcus maripaludis was analyzed. M. maripaludis is a model organism that can be easily genetically modified. Furthermore, its genome has been sequenced, and many of its biochemical pathways have been characterized. FIG. 1. Genetic map of Methanosarcina barkeri ech (A), Methanothermobacter marburgensis ehb (MTH1235-1251) (B), and Methanococcus maripaludis ehb (MMP1631-1629) (C) operons. Genes encoding integral membrane proteins found only in Ehb are indicated in blue, integral ...

Dissimilatory sulphate reduction with acetate as electron donor.

Abstract: Acetate oxidation by sulphate was studied with Desulfobacter postgatei. Cell extracts of the organism were found to contain high activities of the following enzymes: citrate synthase, aconitase, isocitrate dehydrogenase, $\alpha$ -ketoglutarate dehydrogenase, succinate dehydrogenase, fumarase, malate dehydrogenase and pyruvate synthase. It is concluded that acetate oxidation with sulphate in D. postgatei proceeds via the citric acid cycle with the synthesis of pyruvate from acetyl CoA and CO $\_2$ as an anaplerotic reaction. The apparent K $\_S$ for acetate oxidation by D. postgatei as determined in vivo was near 0.2 mM. The apparent K $\_S$ for acetate fermentation to methane and CO $\_2$ by Methanosarcina barkeri was 3 mM. The significantly lower K $_S$ for acetate of the sulphate reducer explains why methane formation from acetate in natural habitats is apparently inhibited by sulphate.

Methanosarcina spelaei sp. nov., a methanogenic archaeon isolated from a floating biofilm of a subsurface sulphurous lake.

Abstract: A novel methanogenic archaeon, strain MC-15T, was isolated from a floating biofilm on a sulphurous subsurface lake in Movile Cave (Mangalia, Romania). Cells were non-motile sarcina-like cocci with a diameter of 2–4 µm, occurring in aggregates. The strain was able to grow autotrophically on H2/CO2. Additionally, acetate, methanol, monomethylamine, dimethylamine and trimethylamine were utilized, but not formate or dimethyl sulfide. Trypticase peptone and yeast extract were not required for growth. Optimal growth was observed at 33 °C, pH 6.5 and a salt concentration of 0.05 M NaCl. The predominant membrane lipids of MC-15T were archaeol and hydroxyarchaeol phosphatidylethanolamine, phosphatidylglycerol, and phosphatidylinositol as well as hydroxyarchaeol phosphatidylserine and archaeol glycosaminyl phosphatidylinositol. The closely related species, Methanosarcina vacuolata and Methanosarcina horonobensis, had a similar composition of major membrane lipids to strain MC-15T. The 16S rRNA gene sequence of strain MC-15T was similar to those of Methanosarcina vacuolata DSM 1232T (sequence similarity 99.3 %), Methanosarcina horonobensis HB-1T (98.8 %), Methanosarcina barkeri DSM 800T (98.7 %) and Methanosarcina siciliae T4/MT (98.4 %). DNA–DNA hybridization revealed 43.3 % relatedness between strain MC-15T and Methanosarcina vacuolata DSM 1232T. The G+C content of the genomic DNA was 39.0 mol%. Based on physiological, phenotypic and genotypic differences, strain MC-15T represents a novel species of the genus Methanosarcina , for which the name Methanosarcina spelaei sp. nov. is proposed. The type strain is MC-15T ( = DSM 26047T = JCM 18469T).

Microbial dehalogenation using methanosarcina

Abstract: A method for bioremediation of hazardous wastes is disclosed. The method can be used for anaerobic treatment of a liquid or slurry hazardous waste stream (e.g., industrial wastewater or sludge) or for treatment of contaminated groundwater. Removal of halogenated (e.g., chlorinated) hydrocarbons, such as tetrachloroethylene, trichloroethylene, 1,1,1-trichloroethane and similar xenobiotics is possible. The method involves biotransformation of (dehalogenation) halogenated hydrocarbons by means of natural methanogenic bacteria, Methanosarcina barkeri strain 227 and/or Methanosarcina vacuolata. These methanogens can accomplish cometabolism of chlorinated hydrocarbons during metabolism of a primary substrate such as hydrogen-carbon dioxide, methanol, methylamine, dimethylamine, trimethylamine and acetate. Reductive dechlorination, energy conservation and control of air pollution are accomplished.

Methanogenesis from Acetate by Methanosarcina barkeri: Catalysis of Acetate Formation from Methyl Iodide, CO2 , and H2 by the Enzyme System Involved

Abstract: Cell suspensions of Methanosarcina barkeri grown on acetate catalyze the formation of methane and CO2 from acetate as well as an isotopic exchange between the carboxyl group of acetate and CO2. Here we report that these cells also mediate the synthesis of acetate from methyl iodide, CO2, and reducing equivalents (H2 or CO), the methyl group of acetate being derived from methyl iodide and the carboxyl group from CO2. Methyl chloride and methyltosylate but not methanol can substitute for methyl iodide in this reaction. Acetate formation from methyl iodide, CO2, and reducing equivalents is coupled with the phosphorylation of ADP. Evidence is presented that methyl iodide is incorporated into the methyl group of acetate via a methyl corrinoid intermediate (deduced from inhibition experiments with propyl iodide) and that CO2 is assimilated into the carboxyl group via a C1 intermediate which does not exchange with free formate or free CO. The effects of protonophores, of the proton-translocating ATPase inhibitor N.N′-di- cyclohexylcarbodiimide, and of arsenate on acetate formation are interpreted to indicate that the reduction of CO2 to the oxidation level of the carboxyl group of acetate requires the presence of an electrochemical proton potential and that acetyl-CoA or acetyl-phosphate rather than free acetate is the immediate product of the condensation reaction. These results are discussed with respect to the mechanism of methanogenesis from acetate.