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B Schink

Publications and source records attributed to B Schink.

At least 73 records · Page 4Linked to original sources
PubMed1994

Fermentation of phenoxyethanol to phenol and acetate by a homoacetogenic bacterium.

A strictly anaerobic gram-positive, rod-shaped bacterium, strain LuPhet1, was isolated from sewage sludge with phenoxyethanol as sole carbon and energy source, and was assigned to the genus Acetobacterium. The new isolate fermented the alkylaryl ether compound phenoxyethanol stoichiometrically to phenol and acetate, whereas phenoxyacetic acid was not degraded. In cell-free extracts of strain LuPhet1, cleavage of the ether linkage was shown, and acetaldehyde was detected as reaction product. Coenzyme A-dependent acetaldehyde: acceptor oxidoreductase, phosphate acetyltransferase, acetate kinase, and carbon monoxide dehydrogenase were measured in cell-free extracts of this strain. Our results indicate that the ether linkage of phenoxyethanol is cleaved by a shift of the hydroxyl group to the subterminal carbon atom, analogous to a corrinoid-dependent diol dehydratase reaction, to form an unstable hemiacetal that releases phenol and acetaldehyde. Obviously, phenoxyethanol is degraded by the same strategy as in anaerobic degradation of the alkyl ether polyethylene glycol.

Acetates↗
PubMed1994

Anaerobic degradation of malonate via malonyl-CoA by Sporomusa malonica, Klebsiella oxytoca, and Rhodobacter capsulatus.

Anaerobic decarboxylation of malonate to acetate was studied with Sporomusa malonica, Klebsiella oxytoca, and Rhodobacter capsulatus. Whereas S. malonica could grow with malonate as sole substrate (Y = 2.0 g.mol-1), malonate decarboxylation by K. oxytoca was coupled with anaerobic growth only in the presence of a cosubstrate, e.g. sucrose or yeast extract (Ys = 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 for K. oxytoca) and hydroxylamine inhibited the enzyme completely, EDTA inhibited partially. In S. malonica and K. 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 proceeds via malonyl-CoA mediated by a CoA transferase and that subsequent decarboxylation to acetyl-CoA is catalyzed, at least with S. malonica and R. capsulatus, by a biotin enzyme.

Acetates↗
PubMed1994

Anaerobic degradation of pimelate by newly isolated denitrifying bacteria.

A C7 dicarboxylic (pimelic) acid derivative is postulated as an intermediate in anaerobic degradation of benzoate. Four strains of Gram-negative, nitrate-reducing bacteria capable of growth with both pimelate and benzoate as sole carbon and energy source were isolated. The metabolism of strain LP-1, which was enriched from activated sludge with pimelate as substrate, was studied in detail. This strain grew only with oxygen or with oxidized nitrogen compounds as electron acceptor. In the presence of nitrate, a wide range of substrates excluding C1 compounds was degraded. The new isolate was catalase- and oxidase-positive, and had one single polar flagellum. Strain LP-1 was tentatively classified within the family Pseudomonadaceae. The catabolism of pimelate and benzoate was studied in cell-free extracts of strain LP-1. Both acids were activated with coenzyme A in a Mg(2E)- and ATP-dependent reaction. The corresponding acyl-CoA synthetases were specifically induced by the respective growth substrate. Pimelate was also activated by CoA transfer from succinyl-CoA. Pimelyl-CoA was oxidized by cell-free extracts in the presence of potassium ferricyanide. Degradation to glutaryl-CoA and acetyl-CoA proceeded by a sequence of beta-oxidation-like reactions. Glutaryl-CoA dehydrogenase and glutaconyl-CoA decarboxylase activities were expressed in cells grown with pimelate or benzoate, indicating the specific involvement of these enzyme activities in anaerobic degradation of these two acids. Enzyme activities responsible for further degradation of the resulting crotonyl-CoA to acetyl-CoA via classical beta-oxidation were also detected.

Anaerobiosis↗
PubMed1994

Anaerobic degradation of catechol by Desulfobacterium sp. strain Cat2 proceeds via carboxylation to protocatechuate.

Under anoxic conditions, most methoxylated mononuclear aromatic compounds are degraded by bacteria, with catechol being formed as an important intermediate. On the basis of our experiments with the sulfate-reducing bacterium Desulfobacterium sp. strain Cat2, we describe for the first time the enzymatic activities involved in the complete anaerobic oxidation of catechol and protocatechuate. Results obtained from experiments with dense cell suspensions of strain Cat2 demonstrated that all enzymes necessary for protocatechuate and benzoate degradation were induced during growth with catechol. In addition, anaerobic oxidation of catechol was found to be a CO2-dependent process. Phenol was not degraded in suspensions of cells grown with catechol. In cell extracts of Desulfobacterium sp. strain Cat2, protocatechuyl-coenzyme A (CoA) was formed from catechol, bicarbonate, and uncombined CoA. This oxygen-sensitive reaction requires high concentrations of both bicarbonate and protein, and only very low levels of enzyme were detected. In a second oxygen-sensitive step, protocatechuyl-CoA was reduced to 3-hydroxybenzoyl-CoA by reductive elimination of the p-hydroxyl group. Further dehydroxylation to benzoyl-CoA was not detectable. Key reactions described for anaerobic degradation of benzoate were catalyzed by cell extracts of strain Cat2, too.

Anaerobiosis↗
PubMed1993

Hydrogen formation from glycolate driven by reversed electron transport in membrane vesicles of a syntrophic glycolate-oxidizing bacterium.

Oxidation of glycolate to 2 CO2 and 3 H2 (delta G degrees' = +36 kJ/mol glycolate) by the proton-reducing, glycolate-fermenting partner bacterium of a syntrophic coculture (strain FlGlyM) depends on a low hydrogen partial pressure (pH2). The first reaction, glycolate oxidation to glyoxylate (E zero' = -92 mV) with protons as electron acceptors (E zero' = -414 mV), is in equilibrium only at a pH2 of 1 microPa which cannot be maintained by the syntrophic partner bacterium Methanospirillum hungatei; energy therefore needs to be spent to drive this reaction. Glycolate dehydrogenase activity (0.3-0.96 U.mg protein-1) was detected which reduced various artificial electron acceptors such as benzyl viologen, methylene blue, dichloroindophenol, K3[Fe(CN)6], and water-soluble quinones. Fractionation of crude cell extract of the glycolate-fermenting bacterium revealed that glycolate dehydrogenase, hydrogenase, and proton-translocating ATPase were membrane-bound. Menaquinones were found as potential electron carriers. Everted membrane vesicles of the glycolate-fermenting bacterium catalyzed ATP-dependent H2 formation from glycolate (30-307 nmol H2.min-1 x mg protein-1). Protonophores, inhibitors of proton-translocating ATPase, and the quinone analog antimycin A inhibited H2 formation from glycolate, indicating the involvement of proton-motive force to drive the endergonic oxidation of glycolate to glyoxylate with concomitant H2 release. This is the first demonstration of a reversed electron transport in syntrophic interspecies hydrogen transfer.

Adenosine Triphosphatases↗
PubMed1992

Energy conservation in fermentative glutarate degradation by the bacterial strain WoG13.

Dicarboxylic acids with 2-5 carbon atoms can be degraded fermentatively by pure cultures of various strictly anaerobic bacteria. The small amount of free energy released in these decarboxylations (about 20-25 kJ mol-1) is conserved as sole source of growth energy either through sodium-pumping decarboxylases or through electrogenic substrate/product transport devices. In the glutarate-fermenting bacterial strain WoG13 a glutaconyl-CoA-decarboxylating enzyme activity was detected. This enzyme was inhibited by avidin and was stimulated by sodium ions. The enzyme activity was partially associated with the cytoplasmic membrane, indicating that energy conservation is accomplished through a sodium-ion-pumping glutaconyl-CoA decarboxylase enzyme.

Bacteria, Anaerobic↗
PubMed1992

Malonate decarboxylase of Malonomonas rubra, a novel type of biotin-containing acetyl enzyme.

Cell suspensions or crude extracts of Malonomonas rubra grown anaerobically on malonate catalyze the decarboxylation of this substrate at a rate of 1.7-2.5 mumol.min-1.mg protein-1 which is consistent with the malonate degradation rate during growth. After fractionation of the cell extract by ultracentrifugation, neither the soluble nor the particulate fraction alone catalyzed the decarboxylation of malonate, but on recombination of the two fractions 87% of the activity of the unfractionated extract was restored. The decarboxylation pathway did not involve the intermediate formation of malonyl-CoA, but decarboxylation proceeded directly with free malonate. The catalytic activity of the enzyme was completely abolished on incubation with hydroxylamine or NaSCN. Approximately 50-65% of the original decarboxylase activity was restored by incubation of the extract with ATP in the presence of acetate, and the extent of reactivation increased after incubation with dithioerythritol. Reactivation of the enzyme was also obtained by chemical acetylation with acetic anhydride. These results indicate modification of the decarboxylase by deacetylation leading to inactivation and by acetylation of the inactivated enzyme specimens leading to reactivation. It is suggested that the catalytic mechanism involves exchange of the enzyme-bound acetyl residues by malonyl residues and subsequent decarboxylation releasing CO2 and regenerating the acetyl-enzyme. The decarboxylase was inhibited by avidin but not by an avidin-biotin complex indicating that biotin is involved in catalysis. A single biotin-containing 120-kDa polypeptide was present in the extract and is a likely component of malonate decarboxylase.

Avidin↗
PubMed1992

Fermentative degradation of glutarate via decarboxylation by newly isolated strictly anaerobic bacteria.

Two strains of new strictly anaerobic, gram-negative bacteria were enriched and isolated from a freshwater (strain WoG13) and a saltwater (strain CuG11) anoxic sediment with glutarate as sole energy source. Strain WoG13 formed spores whereas strain CuG11 did not. Both strains were rod-shaped, motile bacteria growing in carbonate-buffered, sulfide-reduced mineral medium supplemented with 2% of rumen fluid. Both strains fermented glutarate to butyrate, isobutyrate, CO2, and small amounts of acetate. With methylsuccinate, the same products were formed, and succinate was fermented to propionate and CO2. No sugars, amino acids or other organic acids were used as substrates. Molar growth yields (Ys) were very small (0.5-0.9 g cell dry mass/mol dicarboxylate). Cells of strain WoG13 contained no cytochromes, and the DNA base ratio was 49.0 +/- 1.4 mol% guanine-plus-cytosine. Enzyme activities involved in glutarate degradation could be demonstrated in cell-free extracts of strain WoG13. A pathway of glutarate fermentation via decarboxylation of glutaconyl-CoA to crotonyl-CoA is suggested which forms butyrate and partly isobutyrate by subsequent isomerization.

Acetates↗
PubMed1992

A strictly anaerobic nitrate-reducing bacterium growing with resorcinol and other aromatic compounds.

With resorcinol as sole source of energy and organic carbon, two stains of gram-negative, nitrate-reducing bacteria were isolated under strictly anaerobic conditions. Strain LuBRes1 was facultatively anaerobic and catalase- and superoxide dismutase-positive. This strain was affiliated with Alcaligenes denitrificans on the basis of substrate utilization spectrum and peritrichous flagellation. Strain LuFRes1 could grow only under anaerobic conditions with oxidized nitrogen compounds as electron acceptor. Cells were catalase-negative but superoxide dismutase-positive. Since this strain was apparently an obligate nitrate reducer, it could not be grouped with any existing genus. Resorcinol was completely oxidized to CO2 by both strains. Neither an enzyme activity reducing or hydrolyzing the resorcinol molecule, nor an acyl-CoA-synthetase activating resorcylic acids or benzoate was detected in cell-free extracts of cells grown with resorcinol. In dense cell suspensions, both strains produced a compound which was identified as 5-oxo-2-hexenoic acid by mass spectrometric analysis. This would indicate a direct, hydrolytic cleavage of the resorcinol nucleus without initial reduction.

Bacteria, Anaerobic↗
PubMed1992

Energy conservation in malolactic fermentation by Lactobacillus plantarum and Lactobacillus sake.

A comparably poor growth medium containing 0.1% yeast extract as sole non-defined constituent was developed which allowed good reproducible growth of lactic acid bacteria. Of seven different strains of lactic acid bacteria tested, only Lactobacillus plantarum and Lactobacillus sake were found to catalyze stoichiometric conversion of L-malate to L-lactate and CO2 concomitant with growth. The specific growth yield of malate fermentation to lactate at pH 5.0 was 2.0 g and 3.7 g per mol with L. plantarum and L. sake, respectively. Growth in batch cultures depended linearly on the malate concentration provided. Malate was decarboxylated nearly exclusively by the cytoplasmically localized malo-lactic enzyme. No other C4-dicarboxylic acid-decarboxylating enzyme activity could be detected at significant activity in cell-free extracts. In pH-controlled continuous cultures, L. plantarum grew well with glucose as substrate, but not with malate. Addition of lactate to continuous cultures metabolizing glucose or malate decreased cell yields significantly. These results indicate that malo-lactic fermentation by these bacteria can be coupled with energy conservation, and that membrane energetization and ATP synthesis through this metabolic activity are due to malate uptake and/or lactate excretion rather than to an ion-translocating decarboxylase enzyme.

Decarboxylation↗
PubMed1992

Energy conservation by succinate decarboxylation in Veillonella parvula.

Veillonella parvula cannot grow with succinate as sole energy source. However, succinate decarboxylation simultaneous with malate or lactate fermentation increased growth yields by 2.4-3.5 g (mol succinate)-1. Malate was fermented stoichiometrically to acetate and propionate whereas lactate fermentation produced more acetate and considerable amounts of H2. Aspartate was utilized only in the presence of succinate as co-substrate. Methylmalonyl-CoA decarboxylase and ATP-dependent pyruvate carboxylase, but not methylmalonyl-CoA:pyruvate transcarboxylase, were detected in cell-free extracts of malate- or lactate-grown cells. The energetic aspects of these fermentation patterns are discussed.

Acetates↗
PubMed1992

Microbial degradation of natural and of new synthetic polymers.

In landfills, deposited waste material is usually faced with strictly anoxic conditions. This means that the design of new biodegradable polymers must take into consideration that degradation should be possible especially in the absence of molecular oxygen. Poly-beta-hydroxybutyrate is depolymerized by the anaerobic fermenting bacterium Ilyobacter delafieldii through an extracellular hydrolase. Monomers are degraded inside the cells through classical beta-oxidation. Polyalkanoates containing odd-numbered or branched-chain acid monomers should he degraded in an analogous manner; in most cases the final mineralization of these residues requires special pathways. A comparison of the chemistry of natural polymer biodegradation leads to the conclusion that synthetic biodegradable polymers should be designed in the future to contain linkages which can be cleaved by extracellular hydrolytic enzymes. Recent findings on aerobic and anaerobic bacterial degradation of synthetic polyethers suggest that natural evolution of new depolymerizing enzymes, perhaps from existing hydrolases, could be possible in a reasonable amount of time, provided that the monomers are likely energy sources for a broad variety of microbes.

Bacteria, Anaerobic↗
PubMed1992

Reciprocal isomerization of butyrate and isobutyrate by the strictly anaerobic bacterium strain WoG13 and methanogenic isobutyrate degradation by a defined triculture.

Isomerization of butyrate and isobutyrate was investigated with the recently isolated strictly anaerobic bacterium strain WoG13 which ferments glutarate to butyrate, isobutyrate, CO(2), and small amounts of acetate. Dense cell suspensions converted butyrate to isobutyrate and isobutyrate to butyrate. C-nuclear magnetic resonance experiments proved that this isomerization was accomplished by migration of the carboxyl group to the adjacent carbon atom. In cell extracts, both butyrate and isobutyrate were activated to their coenzyme A (CoA) esters by acyl-CoA:acetate CoA-transferases. The reciprocal rearrangement of butyryl-CoA and isobutyryl-CoA was catalyzed by a butyryl-CoA:isobutyryl-CoA mutase which depended strictly on the presence of coenzyme B(12). Isobutyrate was completely degraded via butyrate to acetate and methane by a defined triculture of strain WoG13, Syntrophomonas wolfei, and Methanospirillum hungatei.

Journal Article↗
PubMed1992

Sequential Transhydroxylations Converting Hydroxyhydroquinone to Phloroglucinol in the Strictly Anaerobic, Fermentative Bacterium Pelobacter massiliensis.

The recently isolated fermenting bacterium Pelobacter massiliensis is the only strict anaerobe known to grow on hydroxyhydroquinone (1,2,4-trihydroxybenzene) as the sole source of carbon and energy, converting it to stoichiometric amounts of acetate. In this paper, we report on the enzymatic reactions involved in the conversion of hydroxyhydroquinone and pyrogallol (1,2,3-trihydroxybenzene) to phloroglucinol (1,3,5-trihydroxybenzene). Cell extracts of P. massiliensis transhydroxylate pyrogallol to phloroglucinol after addition of 1,2,3,5-tetrahydroxybenzene (1,2,3,5-TTHB) as cosubstrate in a reaction identical to that found earlier with Pelobacter acidigallici (A. Brune and B. Schink, J. Bacteriol. 172:1070-1076, 1990). Hydroxyhydroquinone conversion to phloroglucinol is initiated in cell extracts without an external addition of cosubstrates. It involves a minimum of three consecutive transhydroxylation reactions characterized by the transient accumulation of two different TTHB isomers. Chemical synthesis of the TTHB intermediates allowed the resolution of the distinct transhydroxylation steps in this sequence. In an initial transhydroxylation, the hydroxyl group in the 1-position of a molecule of hydroxyhydroquinone is transferred to the 5-position of another molecule of hydroxyhydroquinone to give 1,2,4,5-TTHB and resorcinol (1,3-dihydroxybenzene) as products. Following this disproportionation of hydroxyhydroquinone, the 1,2,4,5-isomer is converted to 1,2,3,5-TTHB, an enzymatic activity present only in hydroxyhydroquinone-grown cells. Finally, phloroglucinol is formed from 1,2,3,5-TTHB by transfer of the 2-hydroxyl group to either hydroxyhydroquinone or resorcinol. The resulting coproducts are again cosubstrates in earlier reactions of this sequence. From the spectrum of hydroxybenzenes transhydroxylated by the cell extracts, the minimum structural prerequisites that render a hydroxybenzene a hydroxyl donor or acceptor are deduced.

Journal Article↗
PubMed1992

Enzymes Involved in Anaerobic Polyethylene Glycol Degradation by Pelobacter venetianus and Bacteroides Strain PG1.

In extracts of polyethylene glycol (PEG)-grown cells of the strictly anaerobically fermenting bacterium Pelobacter venetianus, two different enzyme activities were detected, a diol dehydratase and a PEG-degrading enzyme which was characterized as a PEG acetaldehyde lyase. Both enzymes were oxygen sensitive and depended on a reductant, such as titanium citrate or sulfhydryl compounds, for optimal activity. The diol dehydratase was inhibited by various corrinoids (adenosylcobalamin, cyanocobalamin, hydroxocobalamin, and methylcobalamin) by up to 37% at a concentration of 100 muM. Changes in ionic strength and the K ion concentration had only limited effects on this enzyme activity; glycerol inhibited the enzyme by 95%. The PEG-degrading enzyme activity was stimulated by the same corrinoids by up to 80%, exhibited optimal activity in 0.75 M potassium phosphate buffer or in the presence of 4 M KCI, and was only slightly affected by glycerol. Both enzymes were located in the cytoplasmic space. Also, another PEG-degrading bacterium, Bacteroides strain PG1, contained a PEG acetaldehyde lyase activity analogous to the corresponding enzyme of P. venetianus but no diol dehydratase. Our results confirm that corrinoid-influenced PEG degradation analogous to a diol dehydratase reaction is a common strategy among several different strictly anaerobic PEG-degrading bacteria.

Journal Article↗
PubMed1991

Anaerobic degradation of 3-hydroxybenzoate by a newly isolated nitrate-reducing bacterium.

A Gram-negative nitrate-reducing bacterium, strain Asl-3, was isolated from activated sludge with nitrate and 3-hydroxybenzoate as sole source of carbon and energy. The new isolate was facultatively anaerobic, catalase- and oxidase-positive and polarly monotrichously flagellated. In addition to nitrate, nitrite, N2O, and O2 served as electron acceptors. Growth with 3-hydroxybenzoate and nitrate was biphasic: nitrate was completely reduced to nitrite before nitrite reduction to N2 started. Benzoate, 3-hydroxybenzoate, 4-hydroxybenzoate, protocatechuate or phenyl-acetate served as electron and carbon source under aerobic and anaerobic conditions. During growth with excess carbon source, poly-beta-hydroxybutyrate was formed. These characteristics allow the affiliation of strain Asl-3 with the family Pseudomonadaceae. Analogous to the pathway of 4-hydroxybenzoate degradation in other bacteria, the initial step in anaerobic 3-hydroxybenzoate degradation by this organism was activation to 3-hydroxy-benzoyl-CoA in an ATP-consuming reaction. Cell extracts of 3-hydroxybenzoate-grown cells exhibited 3-hydroxybenzoyl-CoA synthetase activity of 190 nmol min-1 mg protein-1 as well as benzoyl-CoA synthetase activity of 86 nmol min-1 mg protein-1. A reductive dehydroxylation of 3-hydroxybenzoyl-CoA could not be demonstrated due to rapid hydrolysis of chemically synthesized 3-hydroxybenzoyl-CoA by cell extracts.

Anaerobiosis↗
PubMed1991

Lithotrophic growth and hydrogen metabolism by Clostridium magnum.

Clostridium magnum, originally described as a non-autotrophic homoacetogenic bacterium, was found to be able to grow with H2/CO2, formate, or methanol with stoichiometric acetate formation, provided that the growth medium contained at least 0.025% (w/v) yeast extract. Hydrogen was also formed as a byproduct of glucose fermentation, and was consumed again after glucose consumption. Hydrogen formation from glucose was independent of growth conditions and reached similar maximal concentrations in mineral media with or without ammonia added as well as in non-growing cultures or in the presence of carbon monoxide.

Carbon Dioxide↗