β-Ketothiolase from <i>Hydrogenomonas eutropha</i> H16 and its significance in the regulation of poly-β-hydroxybutyrate metabolism

Oeding, Volker; Schlegel, H. G.

doi:10.1042/bj1340239

Cited by 243 publications

(116 citation statements)

References 30 publications

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“…Two of these, phaA (H16_A1438; induced twofold) and phaB1 (H16_A1439; induced twofold), are known to encode key enzymes of PHB metabolism, mediating the first two steps of PHB biosynthesis from acetyl-CoA to R-3-hydroxybutyryl-CoA (Schubert et al, 1988;Oeding & Schlegel, 1973) as shown in Fig. 3.…”

Section: Resultsmentioning

confidence: 99%

Genome-wide transcriptome analyses of the ‘Knallgas’ bacterium Ralstonia eutropha H16 with regard to polyhydroxyalkanoate metabolism

et al. 2010

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Ralstonia eutropha H16 is probably the best-studied 'Knallgas' bacterium and producer of poly(3-hydroxybutyrate) (PHB). Genome-wide transcriptome analyses were employed to detect genes that are differentially transcribed during PHB biosynthesis. For this purpose, four transcriptomes from different growth phases of the wild-type H16 and of the two PHB-negative mutants PHB " 4 and DphaC1 were compared: (i) cells from the exponential growth phase with cells that were in transition to stationary growth phase, and (ii) cells from the transition phase with cells from the stationary growth phase of R. eutropha H16, as well as (iii) cells from the transition phase of R. eutropha H16 with those from the transition phase of R. eutropha PHB " 4 and (iv) cells from the transition phase of R. eutropha DphaC1 with those from the transition phase of R. eutropha PHB " 4. Among a large number of genes exhibiting significant changes in transcription level, several genes within the functional class of lipid metabolism were detected. In strain H16, phaP3, accC2, fabZ, fabG and H16_A3307 exhibited a decreased transcription level in the stationary growth phase compared with the transition phase, whereas phaP1, H16_A3311, phaZ2 and phaZ6 were found to be induced in the stationary growth phase. Compared with PHB " 4, we found that phaA, phaB1, paaH1, H16_A3307, phaP3, accC2 and fabG were induced in the wildtype, and phaP1, phaP4, phaZ2 and phaZ6 exhibited an elevated transcription level in PHB " 4. In strain DphaC1, phaA and phaB1 were highly induced compared with PHB " 4. Additionally, the results of this study suggest that mutant strain PHB " 4 is defective in PHB biosynthesis and fatty acid metabolism. A significant downregulation of the two cbb operons in mutant strain PHB " 4 was observed. The putative polyhydroxyalkanoate (PHA) synthase phaC2 identified in strain H16 was further investigated by several functional analyses. Mutant PHB " 4 could be phenotypically complemented by expression of phaC2 from a plasmid; on the other hand, in the mutant H16DphaC1, no PHA production was observed. PhaC2 activity could not be detected in any experiment.

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Section: Resultsmentioning

confidence: 99%

Genome-wide transcriptome analyses of the ‘Knallgas’ bacterium Ralstonia eutropha H16 with regard to polyhydroxyalkanoate metabolism

et al. 2010

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“…The extract of ethanol-grown cells was used as a control. The results indicate that 3-hydroxybutyrate is converted to acetyl-CoA by enzyme reactions reported in Azotobacter beijerinckii (Senior & Dawes, 1973) and Hydrogenomonas eutropha (Oeding & Schlegel, 1973). 3-Hydroxybutyrate is oxidized to acetoacetate by D-3-hydroxybutyrate dehydrogenase (NAD) (0.09 pmol/min/mg protein) which is then activated by acetoacetate : succinate CoA transferase to give acetoacetyl-CoA.…”

Section: The Enzymes Of Ethanol Oxidationmentioning

confidence: 97%

The Oxidation and Assimilation of C2 Compounds by Hyphomicrobium sp.

Attwood

Harder

1974

Journal of General Microbiology

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SUMMARYHyphomicrobium sp. were grown on ethanol, acetate, 3-hydroxybutyrate and methanol. The specific activities of the following enzymes were measured in cellfree extracts of Hyphomicrobium strain x : ethanol dehydrogenase, acetaldehyde dehydrogenase, acetothiokinase, 3-hydroxybutyrate dehydrogenase, b-keto thiolase, citrate synthase, aconitate hydratase, isocitrate dehydrogenase, a-oxoglutarate dehydrogenase, succinate dehydrogenase, fumarate hydratase, malate dehydrogenase, isocitrate lyase, malate synthase, malate dehydrogenase (decarboxylating), phosphoenolpyruvate carboxylase, pyruvate kinase, phosphoenolpyruvate carboxykinase, phosphopyruvate hydratase and phosphoglycerate mutase. From a comparison of the specific activities of these enzymes in the different cell-free extracts, it was concluded that during growth of Hyphomicrobium sp. on either ethanol, acetate or 3-hydroxybutyrate, energy and reducing power is generated mainly by the tricarboxylic acid (TCA) cycle. Carbon is assimilated via the glyoxylate cycle with phosphoenolpyruvate carboxykinase functioning as the main gluconeogenic enzyme. The conversion of ethanol, acetate and 3-hydroxybutyrate to acetylCoA occurs by pathways previously reported in other micro-organisms. I N T R O D U C T I O NStrains of the genus Hyphomicrobium have been isolated and purified from a wide range of natural sources (Attwood & Harder, 1972). When the strains were screened for their ability to grow in mineral media supplemented with different carbon compounds as the sole source of carbon and energy, only a limited number of compounds could be utilized . The C1 compounds methanol, methylamine and formate, the C2 compounds ethanol and acetate, and the C4 compound 3-hydroxybutyrate could support growth. On the basis of these results we decided to investigate the dissimilation and assimilation pathways of (i) C , compounds (Harder, Attwood & Quayle, 1973), and (ii) C2 compounds and 3-hydroxybutyrate. The present paper reports the results obtained when the organisms were grown on ethanol, acetate and 3-hydroxybutyrate. METHODSGrowth of the organism. Hyphomicrobium strains x and G were used throughout the Growth in liquid media was in 3 1 flasks containing 750 ml of an inorganic salts medium investigation. Both strains were maintained as previously reported .

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“…coli cells were suspended in 50 mM phosphate buffer, pH 7.0, and then disrupted by ultrasonication at 4°C. The activities of β-ketothiolase and acetoacetyl-CoA reductase were assayed according to the method of Oeding and Schlegel (1973). PHB synthase activity was determined according to the modified method of Haywood et al (1991).…”

Section: Enzyme Activity Measurementmentioning

confidence: 99%

Molecular and functional analysis of the poly-β-hydroxybutyrate biosynthesis operon of Pseudomonas sp BJ-1

Zhu¹,

Zhang²,

Jiang³

et al. 2010

Genet. Mol. Res.

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ABSTRACT. The operon comprising the genes for poly-β-hydroxybutyrate (PHB) biosynthesis in Pseudomonas sp BJ-1 was cloned and sequenced. Sequence analysis of 8991 bp revealed that the regions contain two related operons. The first operon contains the three genes phbA, phbB and phbC, and the other contains the two genes flp1 and flp2. The deduced amino acid sequences of PHBA and PHBB showed high identity with other bacterial PHB genes. Transcription of the three genes of the first operon is controlled by a single hypothetical promoter region, whereas the other two flp genes are controlled by two hypothetical promoter regions. Analysis of expressed protein at different times showed that PHBA protein levels increased from 0 to 4 h; PHBB and PHBC showed similar kinetics. Detection of enzyme activity showed three proteins with bioactivity and biological function in the synthesis of PHB intermediates.

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β-Ketothiolase from Hydrogenomonas eutropha H16 and its significance in the regulation of poly-β-hydroxybutyrate metabolism

Cited by 243 publications

References 30 publications

Genome-wide transcriptome analyses of the ‘Knallgas’ bacterium Ralstonia eutropha H16 with regard to polyhydroxyalkanoate metabolism

Genome-wide transcriptome analyses of the ‘Knallgas’ bacterium Ralstonia eutropha H16 with regard to polyhydroxyalkanoate metabolism

The Oxidation and Assimilation of C2 Compounds by Hyphomicrobium sp.

Molecular and functional analysis of the poly-β-hydroxybutyrate biosynthesis operon of Pseudomonas sp BJ-1

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