Molecular characterization and properties of (R)-specific enoyl-CoA hydratases from Pseudomonas aeruginosa: metabolic tools for synthesis of polyhydroxyalkanoates via fatty acid ß-oxidation

Tsuge, Takeharu; Taguchi, Kazunori; Seiichi,; Taguchi,; Doi, Yoshiharu

doi:10.1016/s0141-8130(02)00082-x

Cited by 112 publications

(83 citation statements)

References 35 publications

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“…In addition, the -oxidation of dodecanoate generates an abundant amount of acetyl-CoA, which is likely used as a main CoA donor for the PCT reaction. Furthermore, MCL 3HA-CoAs may be supplied from the -oxidation pathway by the introduction of the enoyl-CoA hydratase (phaJ4) gene from Pseudomonas aeruginosa (Tsuge et al, 2003). Thus, the system should meet the required conditions mentioned above.…”

Section: Resultsmentioning

confidence: 99%

Biosynthesis of glycolate-based polyesters containing medium-chain-length 3-hydroxyalkanoates in recombinant Escherichia coli expressing engineered polyhydroxyalkanoate synthase

Matsumoto

Ishiyama

Sakai

et al. 2011

Journal of Biotechnology

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Glycolate(GL)-based polyesters were for the first time produced in the recombinant Escherichia coli fatty acid -oxidation pathway reinforcing mutant LS5218, using extracellularly added GL as a monomer precursor. Cells expressing a Ser325Thr/Gln481Lys mutant of polyhydroxyalkanoate synthase (PhaC1STQK) from Pseudomonas sp. 61-3, propionyl-CoA transferase from Megasphaera elsdenii and enoyl-CoA hydratase from Pseudomonas aeruginosa grown on GL and dodecanoate were found to produce novel copolymers of GL with 3-hydroxyalkanoates (3HAs) (C 4 -C 12 ), P(GL-co-3HA), with a weight-average molecular weight of 34 000. The 1 H NMR analysis of the copolymer revealed the incorporation of GL units into the polymer chain. This result demonstrates that PhaC1STQK polymerized glycolyl-CoA as a monomer substrate. Additionally, the novel lactate(LA)-based polyester P(LA-co-3HA) was produced by substituting GL with LA, indicating that the method is versatile and allows the production of a variety of biopolymers. Keywords:poly (glycolide) poly (lactide) lactate-polymerizing enzyme biobased plastic polyhydroxyalkanoate

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Section: Resultsmentioning

confidence: 99%

Biosynthesis of glycolate-based polyesters containing medium-chain-length 3-hydroxyalkanoates in recombinant Escherichia coli expressing engineered polyhydroxyalkanoate synthase

Matsumoto

Ishiyama

Sakai

et al. 2011

Journal of Biotechnology

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“…The metabolic links between the fatty acid metabolism and PHA biosynthesis are mediated by various enzymes such as enoyl-CoA hydratase (7,8,9,23,34,35), 3-ketoacyl-ACP reductase (22, 27, 33), epimerase (20), and 3-hydroxyacyl-ACP:CoA transacylase (12, 26). The genes encoding these enzymes have been cloned from various bacteria and characterized in detail at molecular level.…”

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confidence: 99%

Identification and Characterization of a New Enoyl Coenzyme A Hydratase Involved in Biosynthesis of Medium-Chain-Length Polyhydroxyalkanoates in Recombinant Escherichia coli

Park

Lee

2003

J Bacteriol

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The biosynthetic pathway of medium-chain-length (MCL) polyhydroxyalkanoates (PHAs) from fatty acids has been established in fadB mutant Escherichia coli strain by expressing the MCL-PHA synthase gene. However, the enzymes that are responsible for the generation of (R)-3-hydroxyacyl coenzyme A (R3HA-CoAs), the substrates for PHA synthase, have not been thoroughly elucidated. Escherichia coli MaoC, which is homologous to Pseudomonas aeruginosa (R)-specific enoyl-CoA hydratase (PhaJ1), was identified and found to be important for PHA biosynthesis in a fadB mutant E. coli strain. When the MCL-PHA synthase gene was introduced, the fadB maoC double-mutant E. coli WB108, which is a derivative of E. coli W3110, accumulated 43% less amount of MCL-PHA from fatty acid compared with the fadB mutant E. coli WB101. The PHA biosynthetic capacity could be restored by plasmid-based expression of the maoC Ec gene in E. coli WB108. Also, E. coli W3110 possessing fully functional ␤-oxidation pathway could produce MCL-PHA from fatty acid by the coexpression of the maoC Ec gene and the MCL-PHA synthase gene. For the enzymatic analysis, MaoC fused with His 6 -Tag at its C-terminal was expressed in E. coli and purified. Enzymatic analysis of tagged MaoC showed that MaoC has enoyl-CoA hydratase activity toward crotonyl-CoA. These results suggest that MaoC is a new enoyl-CoA hydratase involved in supplying (R)-3-hydroxyacyl-CoA from the ␤-oxidation pathway to PHA biosynthetic pathway in the fadB mutant E. coli strain.Polyhydroxyalkanoates (PHAs) are polyesters of (R)-hydroxyalkanoic acids accumulated in numerous bacteria as an energy and carbon storage material under nutrient limiting condition in the presence of excess carbon source (1,17,20). PHAs have been attracting much attention as they can be used as biodegradable polymers (20) and as the sources of chiral pools for the synthesis of fine chemicals (18). The metabolic pathways for the biosynthesis and degradation of PHAs have been well examined in many bacteria (17,20). For example, in short-chain-length-PHA-producing bacteria such as Ralstonia eutropha and Alcaligenes latus, two acetyl coenzyme A (acetylCoA) moieties derived from various carbon sources are condensed to acetoacetyl-CoA by 3-ketothiolase (PhaA) and sequentially converted to (R)-3-hydroxybutyryl-CoA (R3HB-CoA) by acetoacetyl-CoA reductase (PhaB). Then, R3HB-CoA is added to the growing chain of poly(3-hydroxybutyrate) [P(3HB)] by the short-chain-length PHA synthase (PhaC) (29).In pseudomonads belonging to the rRNA homology group I, the intermediates of fatty acid metabolism including enoylCoA, (S)-3-hydroxyacyl-CoA, 3-ketoacyl-CoA, and 3-hydroxyacyl-acyl carrier protein (ACP) are major precursors for medium-chain-length (MCL) PHAs (17,20,36). The metabolic links between the fatty acid metabolism and PHA biosynthesis are mediated by various enzymes such as enoyl-CoA hydratase (7,8,9,23,34,35), 3-ketoacyl-ACP reductase (22, 27, 33), epimerase (20), and 3-hydroxyacyl-ACP:CoA transacylase (12, 26). The genes encoding the...

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“…13) By way of contrast, the three other PhaJs generate medium-chain-length 3HA-CoAs of C6-C12. 7) Such (R)-specific enoyl-CoA hydratase activity toward medium-chain-length enoyl-CoAs of C6-C10 has also been detected in other PHAaccumulating bacteria, including P. oleovorans and P. putida. 14) Although some Pseudomonas strains, including P. stuzeri, 15) P. corrugate, 16) and P. mendocina, 17) generated 3HDD and 3HTD monomers in their PHAs, PhaJ enzymes that hydrate long-chain-length enoylCoAs (more than C12) or unsaturated enoyl-CoAs have yet to be detected in any other PHA-accumulating bacterium.…”

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confidence: 94%

“…6) Additionally, four phaJ genes of Pseudomonas aeruginosa evidence differing specificities for the hydration of trans-2-enoyl-CoA from C4 to C12. 7) However, PhaJ activity toward long-chain-length enoyl-CoAs (more than C12) or unsaturated enoyl-CoAs has not thus far been detected in PHA-producing bacteria.…”

mentioning

confidence: 99%

Overexpression of the (R)-Specific Enoyl-CoA Hydratase Gene fromPseudomonas chlororaphisHS21 inPseudomonasStrains for the Biosynthesis of Polyhydroxyalkanoates of Altered Monomer Composition

Chung

Rhee

2012

Bioscience, Biotechnology, and Biochemistry

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Molecular characterization and properties of (R)-specific enoyl-CoA hydratases from Pseudomonas aeruginosa: metabolic tools for synthesis of polyhydroxyalkanoates via fatty acid ß-oxidation

Cited by 112 publications

References 35 publications

Biosynthesis of glycolate-based polyesters containing medium-chain-length 3-hydroxyalkanoates in recombinant Escherichia coli expressing engineered polyhydroxyalkanoate synthase

Biosynthesis of glycolate-based polyesters containing medium-chain-length 3-hydroxyalkanoates in recombinant Escherichia coli expressing engineered polyhydroxyalkanoate synthase

Identification and Characterization of a New Enoyl Coenzyme A Hydratase Involved in Biosynthesis of Medium-Chain-Length Polyhydroxyalkanoates in Recombinant Escherichia coli

Overexpression of the (R)-Specific Enoyl-CoA Hydratase Gene fromPseudomonas chlororaphisHS21 inPseudomonasStrains for the Biosynthesis of Polyhydroxyalkanoates of Altered Monomer Composition

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