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201.
Forty-four mutants of Alcaligenes eutrophus H 16 were isolated which grew poorly or not at all under autotrophic conditions. Four types were characterized with respect to their defects and their physiological properties. One mutant lacked both enzymes specific for autotrophic CO2 fixation, another one lacked both hydrogenases, and two mutants lacked either the membrane-bound or the soluble hydrogenase. Comparing the results of studies on these mutant types, the following conclusions were drawn: the lack of each hydrogenase enzyme could be partially compensated by the other one; the lack of membrane-bound hydrogenase did not affect autotrophic growth, whereas the lack of the soluble hydrogenase resulted in a decreased autotrophic growth rate. When pyruvate as well as hydrogen were supplied to the wild-type, the cell yield was higher than in the presence of pyruvate alone. Mutant experiments under these conditions indicated that either of both hydrogenases was able to add to the energy supply of the cell. Only the soluble hydrogenase was involved in the control of the rate of hydrogen oxidation by carbon dioxide; the mutant lacking this enzyme did not respond to the presence or absence of CO2. The suppression of growth on fructose by hydrogen could be mediated by either of both hydrogenases alone. 相似文献
202.
Anaerobic degradation of hydroquinone was studied with the fermenting bacterium strain HQGö1. The rate of hydroquinone degradation by dense cell suspensions was dramatically accelerated by addition of NaHCO3. During fermentation of hydroquinone in the presence of 14C-Na2CO3 benzoate was formed as a labelled product, indicating an initial ortho-carboxylation of hydroquinone to gentisate. Gentisate was activated to the corresponding CoA-ester in a CoA ligase reaction at a specific activity of 0.15 mol x min–1 x mg protein–1. Gentisyl-CoA was reduced to benzoyl-CoA with reduced methyl viologen as electron donor by simultaneous reductive elimination of both the ortho and meta hydroxyl group. The specific activity of this novel gentisyl-CoA reductase was 17 nmol x min–1 x mg protein–1. Further degradation to acetate was catalyzed by enzymes which occur also in other bacteria degrading aromatic compounds via benzoyl-CoA. 相似文献
203.
Anaerobic decarboxylation of malonate to acetate was studied withSporomusa malonica, Klebsiella oxytoca, andRhodobacter capsulatus. WhereasS. malonica could grow with malonate as sole substrate (Y=2.0 g·mol–1), malonate decarboxylation byK. oxytoca was coupled with anaerobic growth only in the presence of a cosubstrate, e.g. sucrose or yeast extract (Y
s
=1.1–1.8 g·mol malonate–1).R. capsulatus used malonate anaerobically only in the light, and growth yields with acetate and malonate were identical. Malonate decarboxylation in cell-free extracts of all three bacteria was stimulated by catalytic amounts of malonyl-CoA, acetyl-CoA, or Coenzyme A plus ATP, indicating that actually malonyl-CoA was the substrate of decarboxylation. Less than 5% of malonyl-CoA decarboxylase activity was found associated with the cytoplasmic membrane. Avidin (except forK. oxytoca) and hydroxylamine inhibited the enzyme completely, EDTA inhibited partially. InS. malonica andK. oxytoca, malonyl-CoA decarboxylase was active only after growth with malonate; malonyl-CoA: acetate CoA transferase was found as well. These results indicate that malonate fermentation by these bacteria proceedsvia malonyl-CoA mediated by a CoA transferase and that subsequent decarboxylation to acetyl-CoA is catalyzed, at least withS. malonica andR. capsulatus, by a biotin enzyme.Abbreviations CoASH
Coenzyme A
- EDTA
ethylenediamine tetraacetate 相似文献
204.
The kinetics and specificity of O-demethylation were studied in cell-free extracts of the strictly anaerobic, methanethiol- and dimethylsulfide-producing homoacetogen
Holophaga foetida strain TMBS4 with methanethiol and tetrahydrofolate (H4folate) as methyl acceptors. Extracts of cells grown with 3,4,5-trimethoxybenzoate contained an enzyme system that demethylated
various phenyl methyl ethers with at least one ortho-positioned hydroxyl or methoxyl group (the ortho system) and also contained a decarboxylase. Extracts of cells grown with 3,5-dihydroxyanisole contained an enzyme system
with a novel specificity that demethylated only the meta-hydroxylated compounds 3,5-dihydroxyanisole and 3-hydroxyanisole (the meta system) and lacked a decarboxylase. H4folate-dependent demethylation produced CH3-H4folate. For a photometric in vitro assay of the meta system, the NADPH-consuming phloroglucinol reductase (PR) reaction was coupled to the phloroglucinol-yielding demethylation
of 3,5-dihydroxyanisole. The kinetics of the indicator enzyme PR were studied. The cell extract had a high and stable specific
PR activity. PR was inhibited by phloroglucinol (substrate inhibition) and the substrate analogue 3,5-dihydroxyanisole. Doubling
the PR activity of the coupled enzyme assay by additions of a PR-enriched fraction had no effect, showing that the PR activity
supplied by cell extract did not limit reaction rates. Demethylation activity of the meta system with either methyl acceptor increased with the square of the protein concentration. With H4folate, the in vivo activity could be attained. Kinetic parameters for the methyl acceptors were determined.
Received: 8 November 1996 / Accepted: 13 January 1997 相似文献
205.
From anaerobic digestor sludge of a waste water treatment plant, a gram-negative, strictly anaerobic sulfate-reducing bacterium was isolated with acetone as sole organic substrate. The bacterium was characterized as a new species, Desulfococcus biacutus. The strain grew with acetone with doubling times of 72 h to 120 h; the growth yield was 12.0 (±2.1) g · [mol acetone]-1. Acetone was oxidized completely, and no isopropanol was formed. In labelling studies with 14CO2, cell lipids (including approx. 50% PHB) of acetone-grown cells became labelled 7 times as high as those of 3-hydroxy-buyrate-grown cells. Enzyme studies indicated that acetone was degraded via acetoacetyl-CoA, and that acetone was channeled into the intermediary metabolism after condensation with carbon dioxide to a C4-compound, possibly free acetoacetate. Acetoacetyl-CoA is cleaved by a thiolase reaction to acetyl-CoA which is completely oxidized through the carbon monoxide dehydrogenase pathway. Strain KMRActS was deposited with the Deutsche Sammlung von Mikroorganismen, Braunschweig, under the number DSM 5651. 相似文献
206.
From anoxic sewage sludge a new strictly anaerobic, spore-forming bacterium was isolated with 2-hydroxybutyrate as sole substrate. 2-, 3-, and 4-hydroxybutyrate, 4-chlorobutyrate, crotonate, vinylacetate, and pyruvate were fermented to acetate and butyrate. Fructose was converted to acetate, butyrate, butanol, and H2. Lactate and acrylate were fermented to acetate and propionate. Cells pregrown with lactate fermented 2-hydroxybutyrate to butyrate, propionate and acetate. No inorganic electron acceptors were reduced. The DNA base ratio was 32.0±1.0 mol % and was similar to that of Clostridium propionicum, which was determined to be 35.3±0.5 mol %. Strain LuHBu1 is described as type strain of a new species, Clostridium homopropionicum sp. nov. Another isolate obtained from marine sediment degraded 2-and 3-hydroxybutyrate to acetate and butyrate and was in some respects similar to the known species Ilyobacter polytropus. 相似文献
207.
Identification of two porins in Pelobacter venetianus fermenting high-molecular-mass polyethylene glycols.
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Porins were purified from cells of the anaerobic gram-negative bacterium Pelobacter venetianus grown with 20-kDa polyethylene glycol. After treatment of the cell envelope fraction with sodium dodecyl sulfate-containing solutions, the murein contained only two major peptidoglycan-associated proteins of 14 and 23 kDa. Both proteins were released from the peptidoglycan by the detergent Triton X-100. Genapol X-80 released only the 23-kDa protein. This protein was purified by chromatography on a hydroxyapatite column. It did not form sodium dodecyl sulfate-resistant oligomers. Reconstituted in lipid bilayer membranes, the 23-kDa protein formed cation-selective channels with a single-channel conductance of 230 pS in 1 M KCl. The channel is not a general-diffusion pore, since its conductance depends only moderately on the salt concentration. The channel conducted ammonium much better than potassium or rubidium ions, suggesting that it is probably involved in ammonium uptake. The outer membrane of P. venetianus contains a further, non-murein-associated pore with an unknown molecular mass. It is also cationically selective and has a single-channel conductance of 1.6 nS in 1 M KCl, which suggests that its effective diameter is similar to that of porins from enteric bacteria. 相似文献
208.
The initial reactions involved in anaerobic aniline degradation by the sulfate-reducing Desulfobacterium anilini were studied. Experiments for substrate induction indicated the presence of a common pathway for aniline and 4-aminobenzoate, different from that for degradation of 2-aminobenzoate, 2-hydroxybenzoate, 4-hydroxybenzoate, or phenol. Degradation of aniline by dense cell suspensions depended on CO2 whereas 4-aminobenzoate degradation did not. If acetyl-CoA oxidation was inhibited by cyanide, benzoate accumulated during degradation of aniline or 4-aminobenzoate, indicating an initial carboxylation of aniline to 4-aminobenzoate, and further degradation via benzoate of both substrates. Extracts of alinine or 4-aminobenzoategrown cells activated 4-aminobenzoate to 4-aminobenzoyl-CoA in the presence of CoA, ATP and Mg2+. 4-Aminobenzoyl-CoA-synthetase showed a K
m for 4-aminobenzoate lower than 10 M and an activity of 15.8 nmol · min-1 · mg-1. 4-Aminobenzoyl-CoA was reductively deaminated to benzoyl-CoA by cell extracts in the presence of low-potential electron donors such as titanium citrate or cobalt sepulchrate (2.1 nmol · min-1 · mg-1). Lower activities for the reductive deamination were measured with NADH or NADPH. Reductive deamination was also indicated by benzoate accumulation during 4-aminobenzoate degradation in cell suspensions under sulfate limitation. The results provide evidence that aniline is degraded via carboxylation to 4-aminobenzoate, which is activated to 4-aminobenzoyl-CoA and further metabolized by reductive deamination to benzoyl-CoA. 相似文献
209.
A strictly anaerobic, Gram-positive, rod-shaped bacterium, strain AmMan1, was isolated from freshwater sediment with mandelate (-hydroxy-phenylacetate) as sole carbon and energy source, and was assigned to the genus Acetobacterium. Only the d-enantiomer of mandelate was degraded, and was fermented to acetate and benzoate. Non-aromatic growth substrates (pyruvate, lactate, malate, glycerol, ethylene glycol, and H2/CO2) were fermented to acetate as sole product. Methoxylated aromatics were demethoxylated to the corresponding phenols. The guanine-plus-cytosine content of the DNA was 36.5±1.5%. Carbon monoxide dehydrogenase, dichlorophenol indophenol-reducing lactate dehydrogenase, NAD-dependent mandelate dehydrogenase, phosphate acetyl transferase, acetate kinase, and pyruvate- or phenylglyoxylate-dependent benzylviologen reductase were measured in mandelate-and/or lactate-grown cells, respectively. A pathway of the homoacetogenic fermentation of mandelate is suggested as another example of incomplete substrate oxidation by homoacetogenic bacteria. 相似文献
210.
Three different defined cocultures of glycolatedegrading strictly anaerobic bacteria were isolated from enrichment cultures inoculated with freshwater sediment samples. Each culture contained a primary fermenting bacterium which used only glycolate as growth substrate. These cells were gram-positive, formed terminal oval spores, and did not contain cytochromes. Growth with glycolate was possible only in coculture with either a homoacetogenic bacterium or a hydrogen-utilizing methanogenic bacterium; the overall fermentation balance was either 4 glycolate 3 acetate + 2CO2, or 4 glycolate 3 CH4+5 CO2. These transformations indicate that glycolate was converted by the primary fermenting bacterium entirely to CO2 and reducing equivalents which were transferred to the partner organisms, probably through interspecies hydrogen transfer. The key enzymes of fermentative glycolate degradation were identified in cell-free extracts. An acetyl-CoA and ADP-dependent glyoxylate-converting enzyme activity, malic enzyme, pyruvate synthase, and methyl viologen-dependent hydrogenase were found at comparably high activities suggesting that these bacteria oxidize glycolate through a new pathway via malyl-CoA, and that ATP is synthesized by substrate-level phosphorylation, in a similar manner as found in a recently isolated glyoxylatefermenting anaerobe. 相似文献