Kinetics and mechanism of synthesis and degradation of poly(3-hydroxybutyrate) in Alcaligenes eutrophus

Kawaguchi, Yasushi; Doi, Yoshiharu

doi:10.1021/ma00035a007

Cited by 168 publications

(108 citation statements)

References 7 publications

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“…] 471,000. This value is lower than what is typically observed in a native bacterial PHB producer such as Cupriavidus necator (formerly Alcaligenes eutrophus; Kawaguchi and Doi, 1992). This lower molecular weight may be due to the high levels of PHA synthase observed in our plant lines via western blotting.…”

Section: Discussioncontrasting

confidence: 55%

High Levels of Bioplastic Are Produced in Fertile Transplastomic Tobacco Plants Engineered with a Synthetic Operon for the Production of Polyhydroxybutyrate

et al. 2011

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An optimized genetic construct for plastid transformation of tobacco (Nicotiana tabacum) for the production of the renewable, biodegradable plastic polyhydroxybutyrate (PHB) was designed using an operon extension strategy. Bacterial genes encoding the PHB pathway enzymes were selected for use in this construct based on their similarity to the codon usage and GC content of the tobacco plastome. Regulatory elements with limited homology to the host plastome yet known to yield high levels of plastidial recombinant protein production were used to enhance the expression of the transgenes. A partial transcriptional unit, containing genes of the PHB pathway and a selectable marker gene encoding spectinomycin resistance, was flanked at the 5# end by the host plant's psbA coding sequence and at the 3# end by the host plant's 3# psbA untranslated region. This design allowed insertion of the transgenes into the plastome as an extension of the psbA operon, rendering the addition of a promoter to drive the expression of the transgenes unnecessary. Transformation of the optimized construct into tobacco and subsequent spectinomycin selection of transgenic plants yielded T0 plants that were capable of producing up to 18.8% dry weight PHB in samples of leaf tissue. These plants were fertile and produced viable seed. T1 plants producing up to 17.3% dry weight PHB in samples of leaf tissue and 8.8% dry weight PHB in the total biomass of the plant were also isolated.

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Section: Discussioncontrasting

confidence: 55%

High Levels of Bioplastic Are Produced in Fertile Transplastomic Tobacco Plants Engineered with a Synthetic Operon for the Production of Polyhydroxybutyrate

et al. 2011

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“…5B), in which PhaZd Weu was not detected in the degradation phase. Both the accumulation and degradation of PHB have been reported to occur at the same time under certain conditions in vivo (16 (Table 4). PhaZa1 Weu and PhaZd Weu were found in the PHB inclusion bodies.…”

Section: Discussionmentioning

confidence: 94%

Properties of a Novel Intracellular Poly(3-Hydroxybutyrate) Depolymerase with High Specific Activity (PhaZd) in Wautersia eutropha H16

2005

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“…The propagation of a polyester chain has been proposed to include the formation of an acyl-S enzyme intermediate at two thiol groups and the transesterification to a propagating chain (17). Cys-319 in the A. eutrophus synthase has been demonstrated by mutagenic analysis to play important roles in the catalytic cycles (9), and the corresponding Cys residue is conserved in the PHA synthase of A. caviae at the same position, 319, in a lipase box-like sequence.…”

Section: Resultsmentioning

confidence: 99%

Cloning and analysis of the poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) biosynthesis genes of Aeromonas caviae

1997

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A 5.0-kbp EcoRV-EcoRI restriction fragment was cloned and analyzed from genomic DNA of Aeromonas caviae, a bacterium producing a copolyester of (R)-3-hydroxybutyrate (3HB) and (R)-3-hydroxyhexanoate (3HHx) [P(3HB-co-3HHx)] from alkanoic acids or oils. The nucleotide sequence of this region showed a 1,782-bp poly (3-hydroxyalkanoate) (PHA) synthase gene (phaC Ac [i.e., the phaC gene from A. caviae]) together with four open reading frames (ORF1, -3, -4, and -5) and one putative promoter region. The cloned fragments could not only complement PHA-negative mutants of Alcaligenes eutrophus and Pseudomonas putida, but also confer the ability to synthesize P(3HB-co-3HHx) from octanoate or hexanoate on the mutants' hosts. Furthermore, coexpression of ORF1 and ORF3 genes with phaC Ac in the A. eutrophus mutant resulted in a decrease in the polyester content of the cells. Escherichia coli expressing ORF3 showed (R)-enoyl-coenzyme A (CoA) hydratase activity, suggesting that (R)-3-hydroxyacyl-CoA monomer units are supplied via the (R)-specific hydration of enoyl-CoA in A. caviae. The transconjugant of the A. eutrophus mutant expressing only phaC Ac effectively accumulated P(3HB-co-3HHx) up to 96 wt% of the cellular dry weight from octanoate in one-step cultivation.The utilization of biological systems for production of biodegradable materials is becoming important as a solution of the problems concerning plastic waste and the global environment. Poly(3-hydroxyalkanoates) (PHA) are produced by a wide variety of bacteria as intracellular carbon-and energystorage materials from renewable carbon resources, such as sugars or plant oils (2,6,18,25). Since these bacterial PHA are biodegradable thermoplastics, they have attracted industrial attention as possible candidates for large-scale biotechnological products. At present, more than 90 different monomeric units have been found as constituents of PHA (37).Bacterial PHA can be divided into two groups, depending on the number of carbon atoms in the monomeric units (35). One group of bacteria, including Alcaligenes eutrophus, produces short-chain-length PHA with C 3 -to-C 5 monomer units, while the other group, including Pseudomonas oleovorans, synthesizes medium-chain-length PHA with C 6 -to-C 14 monomer units. Only a few reports are available for bacteria which can synthesize PHA consisting of both short-and medium-chainlength monomer units. For example, Rhodospirillum rubrum (3), Rhodocyclus gelatinosus (19), and Rhodococcus ruber (12) produce terpolymers consisting of C 4 , C 5 , and C 6 3-hydroxyalkanoate (3HA) units from hexanoate, and some pseudomonad strains accumulate PHA consisting of C 4 -to-C 12 3HA units (16,38). Our laboratory has found that a random copolymer of 3-hydroxybutyrate (3HB) and 3-hydroxyhexanoate (3HHx), P(3HB-co-3HHx), is produced by Aeromonas caviae FA440 isolated from soil (7,32). This bacterium synthesizes the copolyester from alkanoic acids of even carbon numbers or from plant oils up to approximately 30 wt% of the cellular dry weight, with a 3HHx fra...

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Kinetics and mechanism of synthesis and degradation of poly(3-hydroxybutyrate) in Alcaligenes eutrophus

Cited by 168 publications

References 7 publications

High Levels of Bioplastic Are Produced in Fertile Transplastomic Tobacco Plants Engineered with a Synthetic Operon for the Production of Polyhydroxybutyrate

High Levels of Bioplastic Are Produced in Fertile Transplastomic Tobacco Plants Engineered with a Synthetic Operon for the Production of Polyhydroxybutyrate

Properties of a Novel Intracellular Poly(3-Hydroxybutyrate) Depolymerase with High Specific Activity (PhaZd) in Wautersia eutropha H16

Cloning and analysis of the poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) biosynthesis genes of Aeromonas caviae

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