scholarly journals The metabolism of protocatechuate by Pseudomonas testosteroni

1968 ◽  
Vol 109 (4) ◽  
pp. 559-568 ◽  
Author(s):  
S. Dagley ◽  
P. J. Geary ◽  
J. M. Wood
Keyword(s):  
Carbon Source ◽  
Cell Extracts ◽  
Lyase Activity ◽  
Water Saturated ◽  
Pseudomonas Testosteroni

1. Protocatechuate 4,5-oxygenase, purified 21-fold from extracts of Pseudomonas testosteroni, was examined in the ultracentrifuge and assigned a mol.wt. of about 140000. 2. When diluted, the enzyme rapidly lost activity during catalysis. Inactivation was partially prevented by l-cysteine. 3. With a saturating concentration of protocatechuate (1·36mm), Km for oxygen was 0·303mm. This value is greater than the concentration of oxygen in water saturated with air at 20°. 4. Cell extracts converted protocatechuate into γ-carboxy-γ-hydroxy-α-oxovalerate, which was isolated as its lactone. 5. γ-Carboxy-γ-hydroxy-α-oxovalerate pyruvate-lyase activity was stimulated by Mg2+ ions and mercaptoethanol. Cells grown with p-hydroxybenzoate as carbon source contained higher concentrations of this enzyme than those grown with succinate.

scholarly journals Effect of Polyphenols on 3-Hydroxy-3-methylglutaryl-Coenzyme A Lyase Activity in Human Hepatoma HepG2 Cell Extracts

2013 ◽  
Vol 36 (12) ◽  
pp. 1902-1906 ◽  
Author(s):  
Saori Nakagawa ◽  
Yuko Kojima ◽  
Koichi Sekino ◽  
Susumu Yamato
Keyword(s):  
Hepg2 Cell ◽  
Coenzyme A ◽  
Human Hepatoma ◽  
Cell Extracts ◽  
Lyase Activity ◽  
Hepatoma Hepg2

scholarly journals The Saccharomyces cerevisiae ICL2 Gene Encodes a Mitochondrial 2-Methylisocitrate Lyase Involved in Propionyl-Coenzyme A Metabolism

2000 ◽  
Vol 182 (24) ◽  
pp. 7007-7013 ◽  
Author(s):  
Marijke A. H. Luttik ◽  
Peter Kötter ◽  
Florian A. Salomons ◽  
Ida J. van der Klei ◽  
Johannes P. van Dijken ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nitrogen Source ◽  
Coenzyme A ◽  
Isocitrate Lyase ◽  
Significant Activity ◽  
Mrna Levels ◽  
Wild Type ◽  
Cell Extracts ◽  
Lyase Activity ◽  
Null Mutants

ABSTRACT The Saccharomyces cerevisiae ICL1 gene encodes isocitrate lyase, an essential enzyme for growth on ethanol and acetate. Previous studies have demonstrated that the highly homologousICL2 gene (YPR006c) is transcribed during the growth of wild-type cells on ethanol. However, even when multiple copies are introduced, ICL2 cannot complement the growth defect oficl1 null mutants. It has therefore been suggested thatICL2 encodes a nonsense mRNA or nonfunctional protein. In the methylcitrate cycle of propionyl-coenzyme A metabolism, 2-methylisocitrate is converted to succinate and pyruvate, a reaction similar to that catalyzed by isocitrate lyase. To investigate whetherICL2 encodes a specific 2-methylisocitrate lyase, isocitrate lyase and 2-methylisocitrate lyase activities were assayed in cell extracts of wild-type S. cerevisiae and of isogenicicl1, icl2, and icl1 icl2 null mutants. Isocitrate lyase activity was absent in icl1 andicl1 icl2 null mutants, whereas in contrast, 2-methylisocitrate lyase activity was detected in the wild type and single icl mutants but not in the icl1 icl2mutant. This demonstrated that ICL2 encodes a specific 2-methylisocitrate lyase and that the ICL1-encoded isocitrate lyase exhibits a low but significant activity with 2-methylisocitrate. Subcellular fractionation studies and experiments with an ICL2-green fluorescent protein fusion demonstrated that theICL2-encoded 2-methylisocitrate lyase is located in the mitochondrial matrix. Similar to that of ICL1, transcription of ICL2 is subject to glucose catabolite repression. In glucose-limited cultures, growth with threonine as a nitrogen source resulted in a ca. threefold induction ofICL2 mRNA levels and of 2-methylisocitrate lyase activity in cell extracts relative to cultures grown with ammonia as the nitrogen source. This is consistent with an involvement of the 2-methylcitrate cycle in threonine catabolism.

Intracellular adenosine triphosphate and cyclic adenosine 3′,5′-monophosphate concentrations during derepression of actinomycin biosynthesis

1982 ◽  
Vol 28 (12) ◽  
pp. 1396-1399 ◽  
Author(s):  
S. Chatterjee ◽  
L. C. Vining
Keyword(s):  
Carbon Source ◽  
Adenosine Triphosphate ◽  
Catabolite Repression ◽  
Antibiotic Production ◽  
Carbon Source Utilization ◽  
Streptomyces Antibioticus ◽  
Cell Extracts

Measurements of adenosine triphosphate and of cyclic adenosine 3′,5′-monophosphate in the mycelium of Streptomyces antibioticus during growth in a medium containing a mixture of glucose and galactose showed no marked changes in concentration during the period when glucose was exhausted and the synthesis of actinomycin began. Thus, these nucleotides do not appear to have a role in mediating catabolite repression of either carbon source utilization or antibiotic production in this organism. Since no NADP-glucohydrolase activity was detected in cell extracts, this enzyme is also excluded from apossible role in regulating actinomycin biosynthesis.

scholarly journals The Pseudomonas aeruginosa devB/SOL Homolog,pgl, Is a Member of the hex Regulon and Encodes 6-Phosphogluconolactonase

2000 ◽  
Vol 182 (14) ◽  
pp. 3934-3941 ◽  
Author(s):  
Paul W. Hager ◽  
M. Worth Calfee ◽  
Paul V. Phibbs
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Carbon Source ◽  
Normal Growth ◽  
Wild Type ◽  
Reading Frame ◽  
Cell Extracts ◽  
Genes Encoding ◽  
Type Rate ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Wild Type Rate

ABSTRACT A cyclic version of the Entner-Doudoroff pathway is used byPseudomonas aeruginosa to metabolize carbohydrates. Genes encoding the enzymes that catabolize intracellular glucose to pyruvate and glyceraldehyde 3-phosphate are coordinately regulated, clustered at 39 min on the chromosome, and collectively form thehex regulon. Within the hex cluster is an open reading frame (ORF) with homology to the devB/SOLfamily of unidentified proteins. This ORF encodes a protein of either 243 or 238 amino acids; it overlaps the 5′ end of zwf (encodes glucose-6-phosphate dehydrogenase) and is followed immediately by eda (encodes the Entner-Doudoroff aldolase). The devB/SOL homolog was inactivated in P. aeruginosa PAO1 by recombination with a suicide plasmid containing an interrupted copy of the gene, creating mutant strain PAO8029. PAO8029 grows at 9% of the wild-type rate using mannitol as the carbon source and at 50% of the wild-type rate using gluconate as the carbon source. Cell extracts of PAO8029 were specifically deficient in 6-phosphogluconolactonase (Pgl) activity. The cloned devB/SOL homolog complemented PAO8029 to restore normal growth on mannitol and gluconate and restored Pgl activity. Hence, we have identified this gene as pgland propose that the devB/SOL family members encode 6-phosphogluconolactonases. Interestingly, three eukaryotic glucose-6-phosphate dehydrogenase (G6PDH) isozymes, from human, rabbit, and Plasmodium falciparum, contain Pgl domains, suggesting that the sequential reactions of G6PDH and Pgl are incorporated in a single protein. 6-Phosphogluconolactonase activity is induced in P. aeruginosa PAO1 by growth on mannitol and repressed by growth on succinate, and it is expressed constitutively in P. aeruginosa PAO8026 (hexR). Taken together, these results establish that Pgl is an essential enzyme of the cyclic Entner-Doudoroff pathway encoded by pgl, a structural gene of the hex regulon.

Degradation of waste 2,4-D residues using a soil bacterium in a sprayer tank system

1991 ◽  
Vol 71 (2) ◽  
pp. 243-246 ◽  
Author(s):  
A. E. Smith ◽  
K. Mortensen
Keyword(s):  
Carbon Source ◽  
Key Words ◽  
Soil Bacterium ◽  
Mineral Salts ◽  
Field Soils ◽  
Tank System ◽  
Pseudomonas Testosteroni

A soil bacterium has been isolated from field soils receiving annual applications of 2,4-D and tentatively identified as Pseudomonas testosteroni Marcus and Talalay. When added to a sprayer tank containing an aerated solution of simple mineral salts and 2,4-D amine formulations, this organism used the herbicide as a carbon source, with stoichiometric release of chloride. This system has been used to biologically degrade 2,4-D amine residues from farm operations and herbicide containers. Key words: Degradation, bacterium, residues, soil, 2,4-D

scholarly journals Conversion of a Thiol Precursor into Aroma Compound 4-mercapto-4-methyl-2-pentanone Using Microbial Cell Extracts

Fermentation ◽  
2021 ◽  
Vol 7 (3) ◽  
pp. 129
Author(s):  
Hao-Kai Li ◽  
Chi-Fong Chang ◽  
Hsuan-Ju Lin ◽  
Jung-Lee Lin ◽  
Yu-Ting Lee ◽  
...  
Keyword(s):  
Colorimetric Method ◽  
Cell Extract ◽  
Microbial Cell ◽  
Aroma Compound ◽  
Resonance Spectroscopy ◽  
Black Currant ◽  
Crude Cell Extract ◽  
Cell Extracts ◽  
Lyase Activity ◽  
Environmentally Friendly Approach

4-Mercapto-4-methyl-2-pentanone (4MMP), a high-impact aroma compound with the box tree and black currant flavors was first identified in wines and could be released by microbial cysteine-S-conjugate β-lyases from its precursors. In this study, various yeasts and bacteria encoding β-lyases were selected to examine their β-lyase activities. A thiol precursor of 4MMP, cysteine-conjugate of 4MMP (cys-4MMP), was synthesized with a purity of >95% in a relatively environmentally friendly approach, and its chemical structure was confirmed by nuclear magnetic resonance spectroscopy. The β-lyase activities of the crude cell extract from the bacteria and yeast strains for different substrates were examined using a colorimetric method. Shewanella putrefaciens cell extract exhibited the highest β-lyase activity for all tested substrates. Additionally, the optimum pH and temperature for their β-lyase activities were determined. To monitor the conversion efficiency of precursor cys-4MMP to 4MMP, liquid chromatography-mass spectrometry was used. Our data indicate that selected bacteria and yeasts could convert cys-4MMP into 4MMP, and S. putrefaciens exhibited the best conversion yield. This study demonstrated the potential use of microbial cell extracts to produce sulfur-containing aroma compounds such as 4MMP.

scholarly journals Physiological Role of β-Phosphoglucomutase inLactococcus lactis

2001 ◽  
Vol 67 (10) ◽  
pp. 4546-4553 ◽  
Author(s):  
Fredrik Levander ◽  
Ulrika Andersson ◽  
Peter Rådström
Keyword(s):  
Growth Rate ◽  
Carbon Source ◽  
Specific Growth Rate ◽  
Type Strain ◽  
Wild Type Strain ◽  
Specific Growth ◽  
Maximum Specific Growth Rate ◽  
Wild Type ◽  
Cell Extracts

ABSTRACT A β-phosphoglucomutase (β-PGM) mutant of Lactococcus lactis subsp. lactis ATCC 19435 was constructed using a minimal integration vector and double-crossover recombination. The mutant and the wild-type strain were grown under controlled conditions with different sugars to elucidate the role of β-PGM in carbohydrate catabolism and anabolism. The mutation did not significantly affect growth, product formation, or cell composition when glucose or lactose was used as the carbon source. With maltose or trehalose as the carbon source the wild-type strain had a maximum specific growth rate of 0.5 h−1, while the deletion of β-PGM resulted in a maximum specific growth rate of 0.05 h−1 on maltose and no growth at all on trehalose. Growth of the mutant strain on maltose resulted in smaller amounts of lactate but more formate, acetate, and ethanol, and approximately 1/10 of the maltose was found as β-glucose 1-phosphate in the medium. Furthermore, the β-PGM mutant cells grown on maltose were considerably larger and accumulated polysaccharides which consisted of α-1,4-bound glucose units. When the cells were grown at a low dilution rate in a glucose and maltose mixture, the wild-type strain exhibited a higher carbohydrate content than when grown at higher growth rates, but still this content was lower than that in the β-PGM mutant. In addition, significant differences in the initial metabolism of maltose and trehalose were found, and cell extracts did not digest free trehalose but only trehalose 6-phosphate, which yielded β-glucose 1-phosphate and glucose 6-phosphate. This demonstrates the presence of a novel enzymatic pathway for trehalose different from that of maltose metabolism in L. lactis.

scholarly journals Halomethane:Bisulfide/Halide Ion Methyltransferase, an Unusual Corrinoid Enzyme of Environmental Significance Isolated from an Aerobic Methylotroph Using Chloromethane as the Sole Carbon Source

1999 ◽  
Vol 65 (10) ◽  
pp. 4301-4312 ◽  
Author(s):  
Catherine Coulter ◽  
John T. G. Hamilton ◽  
W. Colin McRoberts ◽  
Leonid Kulakov ◽  
Michael J. Larkin ◽  
...  
Keyword(s):  
Carbon Source ◽  
Sole Carbon Source ◽  
Cobalt Atom ◽  
Methionine Synthase ◽  
High Yield ◽  
Methylotrophic Bacterium ◽  
Environmental Implications ◽  
16S Rrna Sequence ◽  
Degrading Bacterium ◽  
Cell Extracts

ABSTRACT A novel dehalogenating/transhalogenating enzyme, halomethane:bisulfide/halide ion methyltransferase, has been isolated from the facultatively methylotrophic bacterium strain CC495, which uses chloromethane (CH3Cl) as the sole carbon source. Purification of the enzyme to homogeneity was achieved in high yield by anion-exchange chromatography and gel filtration. The methyltransferase was composed of a 67-kDa protein with a corrinoid-bound cobalt atom. The purified enzyme was inactive but was activated by preincubation with 5 mM dithiothreitol and 0.5 mM CH3Cl; then it catalyzed methyl transfer from CH3Cl, CH3Br, or CH3I to the following acceptor ions (in order of decreasing efficacy): I−, HS−, Cl−, Br−, NO2 −, CN−, and SCN−. Spectral analysis indicated that cobalt in the native enzyme existed as cob(II)alamin, which upon activation was reduced to the cob(I)alamin state and then was oxidized to methyl cob(III)alamin. During catalysis, the enzyme shuttles between the methyl cob(III)alamin and cob(I)alamin states, being alternately demethylated by the acceptor ion and remethylated by halomethane. Mechanistically the methyltransferase shows features in common with cobalamin-dependent methionine synthase from Escherichia coli. However, the failure of specific inhibitors of methionine synthase such as propyl iodide, N2O, and Hg2+to affect the methyltransferase suggests significant differences. During CH3Cl degradation by strain CC495, the physiological acceptor ion for the enzyme is probably HS−, a hypothesis supported by the detection in cell extracts of methanethiol oxidase and formaldehyde dehydrogenase activities which provide a metabolic route to formate. 16S rRNA sequence analysis indicated that strain CC495 clusters with Rhizobium spp. in the alpha subdivision of the Proteobacteria and is closely related to strain IMB-1, a recently isolated CH3Br-degrading bacterium (T. L. Connell Hancock, A. M. Costello, M. E. Lidstrom, and R. S. Oremland, Appl. Environ. Microbiol. 64:2899–2905, 1998). The presence of this methyltransferase in bacterial populations in soil and sediments, if widespread, has important environmental implications.

Isocitrate lyase in Coprinus lagopus (sensu Buller)

1969 ◽  
Vol 15 (6) ◽  
pp. 637-640 ◽  
Author(s):  
P. J. Casselton ◽  
M. O. Fawole ◽  
L. A. Casselton
Keyword(s):  
Carbon Source ◽  
Isocitrate Lyase ◽  
Glucose Medium ◽  
Lyase Activity ◽  
Acetate Medium

Isocitrate lyase activity was detected in cell-free extracts of both a monokaryon and a dikaryon of Coprinus lagopus sensu Buller grown in media containing either acetate or glucose as carbon source. The activity in extracts of mycelium grown in an acetate medium was greater than that in extracts of mycelium grown in a glucose medium but marked differences between extracts of monokaryons and dikaryons grown in similar media were not observed. The isocitrate lyase activity could be largely sedimented by centrifugation at 10 000 × g for 20 minutes.

scholarly journals l-Malyl-Coenzyme A/β-Methylmalyl-Coenzyme A Lyase Is Involved in Acetate Assimilation of the Isocitrate Lyase-Negative Bacterium Rhodobacter capsulatus

2005 ◽  
Vol 187 (4) ◽  
pp. 1415-1425 ◽  
Author(s):  
Michael Meister ◽  
Stephan Saum ◽  
Birgit E. Alber ◽  
Georg Fuchs
Keyword(s):  
Rhodobacter Capsulatus ◽  
Coenzyme A ◽  
Isocitrate Lyase ◽  
Glyoxylate Cycle ◽  
Gene Clusters ◽  
Malate Synthase ◽  
Acetate Metabolism ◽  
Acetate Assimilation ◽  
Cell Extracts ◽  
Lyase Activity

ABSTRACT Cell extracts of Rhodobacter capsulatus grown on acetate contained an apparent malate synthase activity but lacked isocitrate lyase activity. Therefore, R. capsulatus cannot use the glyoxylate cycle for acetate assimilation, and a different pathway must exist. It is shown that the apparent malate synthase activity is due to the combination of a malyl-coenzyme A (CoA) lyase and a malyl-CoA-hydrolyzing enzyme. Malyl-CoA lyase activity was 20-fold up-regulated in acetate-grown cells versus glucose-grown cells. Malyl-CoA lyase was purified 250-fold with a recovery of 6%. The enzyme catalyzed not only the reversible condensation of glyoxylate and acetyl-CoA to l-malyl-CoA but also the reversible condensation of glyoxylate and propionyl-CoA to β-methylmalyl-CoA. Enzyme activity was stimulated by divalent ions with preference for Mn2+ and was inhibited by EDTA. The N-terminal amino acid sequence was determined, and a corresponding gene coding for a 34.2-kDa protein was identified and designated mcl1. The native molecular mass of the purified protein was 195 ± 20 kDa, indicating a homohexameric composition. A homologous mcl1 gene was found in the genomes of the isocitrate lyase-negative bacteria Rhodobacter sphaeroides and Rhodospirillum rubrum in similar genomic environments. For Streptomyces coelicolor and Methylobacterium extorquens, mcl1 homologs are located within gene clusters implicated in acetate metabolism. We therefore propose that l-malyl-CoA/β-methylmalyl-CoA lyase encoded by mcl1 is involved in acetate assimilation by R. capsulatus and possibly other glyoxylate cycle-negative bacteria.

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