Showing papers on "Methanosarcina barkeri published in 1983"

Distribution of polyamines in methanogenic bacteria.

Abstract: Members of all four families of methanogenic bacteria were analyzed for polyamine concentrations. High-performance liquid chromatography analysis of dansylated cell extracts revealed typical polyamine patterns for each family. Members of Methanobacteriaceae (family I) were characterized by very low polyamine concentrations; members of Methanococcaceae (family II) were characterized by putrescine and high spermidine concentrations; members of Methanomicrobiaceae (family III) were characterized by the presence of putrescine, spermidine, and sym-homospermidine; and members of Methanosarcinaceae (family IV) contained only high concentrations of sym-homospermidine in addition to putrescine. The highest polyamine concentration was found in Methanosarcina barkeri Julich, with 0.35% putrescine in the dry cell material. The polyamine distribution found coincides with the dendrogram based on comparative cataloguing of 16S rRNA and offers a new, rapid chemotaxonomic method for characterizing methanogenic bacteria. Variation of the growth substrates (H2-CO2, methanol, acetate, and trimethylamine) for M. barkeri resulted in quantitative but not qualitative differences in polyamine composition.

Methyltransferases involved in methanol conversion by Methanosarcina barkeri.

Abstract: 2-(Methylthio)ethanesulfonate (CH3S-CoM) is formed as an intermediate in methanogenesis from methanol by cell-free extracts of Methanosarcina barkeri. The enzyme system involved in the methyltransfer from methanol to 2-mercaptoethanesulfonate (HS-CoM) was resolved into two enzyme fractions. One enzyme (methanol: 5-hydroxybenzimidazolylcobamide methyltransferase) appears to be a cobalamin-containing protein, which is oxygen sensitive. The other enzyme (Co-methyl-5-hydroxybenzimidazolylcobamide: HS-CoM methyltransferase) was purified. It is insensitive to oxygen and it transfers also the methylgroup from Co-methyl-5,6-dimethylbenzimidazolylcobamide to HS-CoM.

Energetics of Growth of a Defined Mixed Culture of Desulfovibrio vulgaris and Methanosarcina barkeri: Interspecies Hydrogen Transfer in Batch and Continuous Cultures

Abstract: Interspecies hydrogen transfer was studied in Desulfovibrio vulgaris-Methanosarcina barkeri mixed cultures. Experiments were performed under batch and continuous growth culture conditions. Lactate or pyruvate was used as an energy source. In batch culture and after 30 days of simultaneous incubation, these organisms were found to yield 1.5 mol of methane and 1.5 mol of carbon dioxide per mol of lactate fermented. When M. barkeri served as the hydrogen acceptor, growth yields of D. vulgaris were higher compared with those obtained on pyruvate without any electron acceptor other than protons. In continuous culture, all of the carbon derived from the oxidation of lactate was recovered as methane and carbon dioxide, provided the dilution rate was minimal. Increasing the dilution rate induced a gradual accumulation of acetate, causing acetate metabolism to cease at above μ = 0.05 h−1. Under these conditions all of the methane produced originated from carbon dioxide. The growth yields of D. vulgaris were measured when sulfate or M. barkeri was the electron acceptor. Two key observations resulted from the present study. First, although sulfate was substituted by M. barkeri, metabolism of D. vulgaris was only slightly modified. The coculture-fermented lactate produced equimolar quantities of carbon dioxide and methane. Second, acetogenesis and methane formation from acetate were completely separable.

Betaine: New Oxidant in the Stickland Reaction and Methanogenesis from Betaine and l-Alanine by a Clostridium sporogenes-Methanosarcina barkeri Coculture

Abstract: Growing and nongrowing cells of Clostridium sporogenes fermented betaine with l-alanine, l-valine, l-leucine, and l-isoleucine as electron donors in a coupled oxidation-reduction reaction (Stickland reaction). For the substrate combinations betaine and l-alanine and betaine and l-valine balance studies were performed; the results were in agreement with the following fermentation equation: 1 R- CH(NH2)-COOH + 2 betaine + 2 H2O → 1 R-COOH + 1 CO2 + 1 NH3 + 2 trimethylamine + 2 acetate. Growth and production of trimethylamine were strictly dependent on the presence of selenite in the medium. With cell suspensions it was shown that C. sporogenes was unable to catabolize betaine as a single substrate. Betaine, however, was reduced to trimethylamine and acetate under an atmosphere of molecular hydrogen. For the reduction of betaine by cell extracts of C. sporogenes, dimercaptans such as 1,4-dithiothreitol could serve as electron donors. No betaine reductase activity was detected in cells grown in a complex medium without betaine. The pH optimum of betaine reductase was at pH 7.3. When C. sporogenes was cocultured with Methanosarcina barkeri strain Fusaro on betaine together with l-alanine, an almost complete conversion of the two substrates to CH4, NH3, and presumably CO2 was observed.

Activation and inactivation of methanol: 2-mercaptoethanesulfonic acid methyltransferase from Methanosarcina barkeri.

Abstract: Methanol is converted to methane by crude extracts of Methanosarcina barkeri. The first reaction involved in this process, is catalyzed by methanol:2-mercaptoethanesulfonic acid methyltransferase (EC 2.1.1.-). The methyltransferase has an optimum at pH 6.5 and is not inhibited by 2-bromoethanesulfonic acid. Pyridoxal-5'-phosphate acts as an inhibitor (Ki = 0.30 mM). The methyltransferase was tested in the presence of 2-bromoethanesulfonic acid, which inhibits the conversion of 2-(methylthio)ethanesulfonic acid to methane. The reaction is subject to activation and inactivation. Inactivation is brought about by the presence of oxygen, flavin mononucleotide, flavin adenine dinucleotide, and 2-(methylthio)ethanesulfonic acid, the product of the reaction. Activation of the system requires the presence of ATP and Mg2+ and of hydrogen. Hydrogen can be replaced by enzymatic systems, such as pyruvate dehydrogenase, which deliver free hydrogen.

Continuous Anaerobic Digestion with Methanosarcina barkeri

Abstract: In spite of the increasing importance of anaerobic digestion, little is known about the kinetic data of relevant microorganisms. In order to obtain a dependable layout of waste water reactors and in order to identify limiting facrors, it is essential to evaluate such kinetic data by means of appropriate measurements. Such experiments should be carried out continuously in a chemostat in order to establish reproducible conditions.’ A pure culture should be used, so that besides the reaction conditions, the “catalyst conditions” are also well defined.’ Acetic acid is a suitable substrate since the methanation of acetic acid is thought to be the limiting step during the anaerobic digestion of many different residues. Acetic acid itself causes chemical oxygen demand (COD) in the waste water of important industrial processes, for example, in the paper and pulp industry.’ Here in the condensate from the concentration of the spent sulfite liquor, acetic acid is found as almost the only carbon source. Such “distilled” waste water is very suitable for the transfer of results with model waste water to a case of practical importance. Furthermore, since only very few microorganisms can grow anaerobically with acetic acid as the only carbon source, such a culture is potentially autosterile. Methanosarcina barkeri, was chosen as a “catalyst” due to the frequent occurrence of this strain in many digestion plants.

Ultrastructural investigation of 12 Methanosarcinae and related species grown on methanol for occurrence of polyphosphatelike inclusions

Abstract: A comparative investigation by electron microscopy of 12 strains and species belonging to the Methanosarcinaceae family of methanogenic bacteria revealed the existence of polyphosphatelike bodies during growth on methanol in an optimized medium. The Methanosarcina barkeri strains Fusaro, 227, and Wiesmoor, and M. vacuolata were found to contain smoothly marginated, polyphosphatelike inclusions. They were identified by a positive Neisser stain, by the absence of a boundary membrane, and by X-ray microanalysis in situ. They had a diameter of 0.15–0.25 µm and contained the elements Ca, P, and Fe, and sometimes Mg, S, and Cl. In addition to the polyphosphatelike bodies, coarse-looking electron-dense "granula" were observed in the cytoplasm of most of the strains. They had an average diameter of 30 nm and the same elemental composition as the polyphosphatelike bodies. As demonstrated for M. barkeri Fusaro, the "granula" were also present in cells grown in acetate and H2–CO2.

Methanogenesis from Choline by a Coculture of Desulfovibrio sp. and Methanosarcina barkeri.

Abstract: A sulfate-reducing vibrio was isolated from a methanogenic enrichment with choline as the sole added organic substrate. This organism was identified as a member of the genus Desulfovibrio and was designated Desulfovibrio strain G1. In a defined medium devoid of sulfate, a pure culture of Desulfovibrio strain G1 fermented choline to trimethylamine, acetate, and ethanol. In the presence of sulfate, more acetate and less ethanol were formed from choline than in the absence of sulfate. When grown in a medium containing sulfate, a coculture of Desulfovibrio strain G1 and Methanosarcina barkeri strain Fusaro degraded choline almost completely to methane, ammonia, and hydrogen sulfide and presumably to carbon dioxide. Methanogenesis occurred in two distinct phases separated by a lag of about 6 days. During the first phase of methanogenesis choline was completely converted to trimethylamine, acetate, hydrogen sulfide, and traces of ethanol by the desulfovibrio. M. barkeri fermented trimethylamine to methane, ammonia, and presumably carbon dioxide via dimethyl- and methylamine as intermediates. Simultaneously, about 60% of the acetate expected was metabolized. In the second phase of methanogenesis, the residual acetate was almost completely catabolized.

Radioassay for Hydrogenase Activity in Viable Cells and Documentation of Aerobic Hydrogen-Consuming Bacteria Living in Extreme Environments

Abstract: An isotopic tracer assay based on the hydrogenase-dependent formation of tritiated water from tritium gas was developed for in life analysis of microbial hydrogen transformation. This method allowed detection of bacterial hydrogen metabolism in pure cultures or in natural samples obtained from aquatic ecosystems. A differentiation between chemical-biological and aerobic-anaerobic hydrogen metabolism was established by variation of the experimental incubation temperature or by addition of selective inhibitors. Hydrogenase activity was shown to be proportional to the consumption or production of hydrogen by cultures of Desulfovibrio vulgaris, Clostridium pasteurianum, and Methanosarcina barkeri. This method was applied, in connection with measurements of free hydrogen and most-probable-number enumerations, in aerobic natural source waters to establish the activity and document the ecology of hydrogen-consuming bacteria in extreme acid, thermal, or saline environments. The utility of the assay is based in part on the ability to quantify bacterial hydrogen transformation at natural hydrogen partial pressures, without the use of artificial electron acceptors.

Propionate assimilation by methanogenic bacteria

Abstract: Methanobacterium thermoautotrophicum, Methanobrevibacter arboriphilus, and Methanosarcina barkeri were found to assimilate propionate when growing on media supplemented with this volatile fatty acid. [1-14C]propionate was almost exclusively incorporated into isoleucine, only C-2 of which became labelled. Assimilation of propionate by M. thermoautotrophicum was specifically inhibited by isoleucine, by 2-methylbutyrate, and by 2-oxobutyrate, whereas there was little or no effect by leucine, valine, butyrate, and acetate. This finding indicates that propionate assimilation is under regulatory control by intermediates and/or the product of isoleucine biosynthesis.

Energetics of growth of a defined mixed culture of Desulfovibrio vulgaris and Methanosarcina barkeri: maintenance energy coefficient of the sulfate-reducing organism in the absence and presence of its partner.

Abstract: The maintenance energy coefficient of Desulfovibrio vulgaris was studied by using a chemostat, with Methanosarcina barkeri or sulfate as the electron acceptor; lithium lactate or sodium pyruvate served as the electron donor. The experiments showed that the growth energetics of D. vulgaris or M. barkeri were greatly affected by maintenance energy coefficients. When D. vulgaris grew on lactate or pyruvate medium with sulfate, these coefficients reached 4.40 and 2.80 mM g-1 h-1, respectively; on lactate medium in the presence of M. barkeri the same coefficient reached a value of 2.90 mM g-1 h-1. Results also showed that the increase of the value of the maintenance energy coefficient corresponded to a decrease of the biomass produced. D. vulgaris maximal growth yield values calculated by use of the Pirt equation were slightly higher with M. barkeri (maximal growth yield, 10 g/mol) than with sulfate (maximal growth yield, 7.5 g/mol). This finding could be interpreted by reference to the ATP-generating reactions involved in D. vulgaris growth in the presence of sulfate or M. barkeri.

Energy-dispersive X-ray microanalysis of the methanogenMethanosarcina barkeri ‘Fusaro’ grown on methanol and in the presence of heavy metals

Abstract: The general distribution of metals in cell compartments ofMethanosarcina barkeri ‘Fusaro’ was studied by energy-dispersive X-ray microanalysis. Cells were cultivated on methanol, acetate, or on a mixture of H2+CO2 as substrate. At decreasing degree (polyphosphate bodies ≫ clusters of small granula>cell walls, inside a cell clump> cytoplasma), the elemental pattern of Ca, P, and Fe could be seen in all cell compartments. Cells grown on methanol were further investigated for their ability to incorporate added heavy metal ions. After completed growth and methane production, Pb or Cd was found to a considerable extent in all cell compartments, whereas Zn or Cu was incorporated only to a minor degree. Evidence for an uptake of Hg could not be obtained.

Effect of some physical and chemical parameters on the fermentation of cellulose to methane by a coculture system

Abstract: In the fermentation of cellulose to methane by the triculture of Acetivibrio cellulolyticus – Desulfovibrio sp. – Methanosarcina barkeri, methanogenesis was the rate-limiting step. The optimal temperature was 35 °C. In the presence of initially added hydrogen, initiation of cellulose hydrolysis was delayed until most of the hydrogen was metabolized, and then the fermentation which followed was comparable with the N2: CO2 control. Increased CH4 yields and rates of formation were stoichiometrically related to the utilization of the hydrogen initially present. Added acetate had no effect on cellulolysis. The increased yields of CH4 observed could be accounted for by utilization of the added acetate. When Methanobrevibacter sp. was included in the coculture (without added H2 or acetate), no hydrogen accumulated and lower rates of acetate utilization and subsequent CH4 evolution were observed. These results suggest a requirement for hydrogen by M. barkeri for efficient acetate utilization. Controlling the pH a...

The Utilization of Inorganic Pyrophosphate, Tripolyphosphate, and Tetrapolyphosphate as Energy Sources for the Growth of Anaerobic Bacteria

Abstract: Inorganic pyrophosphate (PPi) has been shown to be utilized as a source of energy for the growth of a number of pure cultures of anaerobic bacteria and isolates from marine and fresh water environments. The microorganisms generally require a source of fixed carbon such as yeast extract for growth but with some of the sulfate reducing bacteria, acetate and sulfate were also required. Growth, as determined by optical density, cell numbers, or protein, is proportional to the concentration of PPi over a limited range and growth is always accompanied by the hydrolysis of PPi to orthophosphate. PPi was also demonstrated to modify fermentations without effecting growth Thus, methane formation by Methanosarcina barkeri from acetate, methanol or H2 plus CO2 is stimulated 2–3 fold in the presence of PPi and, with the latter two substrates, an accumulation of acetate is observed without a significant increase in growth. Enrichment cultures have been obtained using tripolyphosphate and tetrapolyphosphate and it is evident that these polyphosphates can also be utilized as a source of energy for the growth of anaerobic microorganisms. An enzyme has been purified from Thermoanaerobacter ethanolicus which exhibits both PPi:acetate kinase and ATP:acetate kinase activities and it is suggested that this kinase may be responsible for the widespread ability among anaerobic bacteria to utilize PPi as a source of energy.

Formation of ethylamine and methane from dimethylethylamine by Methanosarcina barkeri

Abstract: Compounds such as tri-, diand monomethylamine have been shown to serve as growth substrates for Methanosarc ina barkeri [1,2]. The products formed from these compounds are methane, carbon dioxide and ammonia. Dimethylethylamine has also been reported to serve as growth substrate for M. barkeri [1], but its fate in this fermentation has not been studied. Here we show that both methyl groups are removed from that compound during methanogenesis and that ethylamine is a product.

Measurement of number of mold and methane formation activity of methanosarcina barkeri

Abstract: PURPOSE: To measure the number of molds of Methanosarcina barkeri in a methane fermentation tank, etc. of a sewage treatment device without obstruction of coexisting admixtures, by measuring the intensity of fluorescence radiated from a specimen which is irradiated with excitation rays in a specific wavelength range. CONSTITUTION: The light rays from the light source 13 are introduced from the light filter 12 through the selector 11, the condenser 10, and the optical fiber 9 to the interior of the methane fermentation tank 8, a specimen is irradiated with the light rays, fluorescence radiated from the specimen is sent from the optical fiber 9 through the condenser 15, and the opticall filter 16 to the phototube 17, and the intensity of the light rays is measured by the detecting part 19. The prepared value as a parameter is used to obtain the number of molds and methane formation activity of Methanosarcina barkeri. In the operation, (i) light rays in a wavelength range of 220W255nm, 260W305nm, are used as the excitation rays (selected by the light filter 12), and (ii) light rays in a wavelength range of 330W370nm as the fluorescence (selected by the light filter 16). COPYRIGHT: (C)1984,JPO&Japio

Distribution ofPolyamines inMethanogenic Bacteria

Abstract: in addition toputrescine. Thehighest polyamine concentration wasfoundin Methanosarcina barkeri Julich, with0.35%putrescine inthedrycell material. Thepolyamine distribution foundcoincides withthedendrogram basedon comparative cataloguing of16SrRNAandoffers anew,rapid chemotaxonomic method forcharacterizing methanogenic bacteria. Variation ofthegrowth substrates (H2-CO2, methanol, acetate, andtrimethylamine) forM.barkeri resulted inquantitative butnotqualitative differences inpolyamine composition.