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PubMed2003

Recovery of poly(3-hydroxybutyrate) from high cell density culture of Ralstonia eutropha by direct addition of sodium dodecyl sulfate.

A simple and effective method for the recovery poly(3-hydroxybutyrate) [P(3HB)] directly from high cell density culture broth with no pretreatment steps has been developed. This method consists of direct addition of sodium dodecyl sulfate (SDS) to the culture broth, shaking, heat treatment, and washing steps. When the SDS/biomass ratio was higher than 0.4, the purity of recovered P(3HB) was over 95% for various cell concentrations of 50-300 g dry cell l(-1), with the highest value of 97%. The recovery of P(3HB) was over 90% regardless of cell concentration and SDS dosage (SDS/biomass ratios, 0.1-0.7). One g SDS digests 0.72 g non-P(3HB) cell materials. The reduction in molecular weight, due to degradation of P(3HB) by SDS, was negligible.

Bioreactors↗
PubMed2003

Enhanced biosynthesis of polyhydroxyalkanoates in a mutant strain of Rhizobium meliloti.

Strains of Rhizobium spp. isolated from leguminous plants and standard strains accumulated 27% to 57% polyhydroxyalkanoate (PHA) of their cell biomass. Among these cultures, one strain of Rhizobium meliloti synthesized 10-30% more PHA than others and contained 3% hydroxyvalerate (HV) when grown on sucrose as carbon substrate. The occurrence of hydroxybutyrate (HB) and HV was confirmed by GC and 1H NMR analysis. Treatment of the culture with 4'-N-piperidinobutyl-2-chlorophenoxazine resulted in a mutant which synthesized up to 69%, PHA of the cell biomass with an improved yield of 11 to 47% under different carbon and nitrogen ratios, compared to the parent strain.

Carbon↗
PubMed2003

Production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with high molar fractions of 3-hydroxyvalerate by a threonine-overproducing mutant of Alcaligenes sp. SH-69.

A threonine overproducing mutant of Alcaligenes sp. SH-69 was isolated and its ability to produce poly(3-hydroxybutyrate-co-3-hydroxyvalerate), poly(3HB-co-3HV), was investigated. The 3HV fraction in poly(3HB-co-3HV) produced from glucose as the sole carbon source exceeded 22 mol%, which is approximately six times higher than that achieved by the wild type under the same culture conditions. Furthermore, the addition of a relatively low concentration (10 mM) of propionic acid, valeric acid or levulinic acid to the glucose medium greatly increased the molar fraction of 3HV in the copolyester, to 38-77 mol%. The results suggest that metabolic engineering of the biosynthetic pathways supplying polyhydroxyalkanoate monomers, such as the threonine biosynthetic pathway, can lead to new poly(3HB-co-3HV)-producing strains.

Alcaligenes↗
PubMed2003

Poly(3-hydroxybutyrate) synthesis in fed-batch culture of Ralstonia eutropha with phosphate limitation under different glucose concentrations.

Poly(3-hydroxybutyrate) (PHB) was produced by fed-batch cultures of Ralstonia eutropha with phosphate limitation under different glucose concentrations. When glucose was kept at 2.5 g l(-1), cell growth and PHB synthesis were limited due to the shortage of carbon source but a higher PHB content occurred in the cell-growth stage. This shows that a low glucose concentration is favorable for PHB accumulation in R. eutropha. PHB obtained with glucose at 9 g l(-1) is 1.6 times that obtained with 40 g l(-1). When glucose was in the range of 9 to 40 g l(-1), PHB concentration and productivity decreased significantly with the increase of glucose concentration. The highest PHB productivity was obtained with glucose at 9 g l(-1).

Bioreactors↗
PubMed2003

Functional role of granule-associated genes, phaP and phaR, in poly-beta-hydroxybutyrate biosynthesis in recombinant E. coli harboring phbCAB operon.

The expression characteristic of two granule-associated genes, phaP and phaR, in a recombinant E. coli harboring the phbCAB operon was investigated. Polybetamydroxybutyrate (PHB) accumulation increased up from 16% to 57% (w/w) after transformation of the granule-associated genes due to the stabilization of PHB granules rather than by a direct effect on PHB biosynthetic enzymes. The morphology of PHB granules also varied depending on the transformed phaP and phaR genes.

Bacterial Proteins↗
PubMed2000

Fine structure of fossilized bacteria in Volyn kerite.

Ultrathin sectioning and cryofracture of fibrous kerite, sampled from 1.8-1.75 billion year old Volyn sediments (Ukraine), revealed in bacteria-like bodies the presence of structures similar to sheath, cell wall, periplasm, cytoplasm, septum, membranes, intramembrane particles, poly-beta-hydroxybutyrate inclusions. On the strength of these data and also the fatty acid profiles of these microfossils, we concluded that fibrous kerites are biogenic formations, namely fossilized bacterial mats.

Bacteria↗
PubMed2004

Microbial degradation of poly(D-3-hydroxybutyrate) by a new thermophilic streptomyces isolate.

A new thermophilic microorganism capable of degrading poly(D-3-hydroxybutyrate) (PHB) was isolated from soil. A phylogenetic analysis based on 16S rDNA sequences indicated that the new isolate belongs to genus Streptomyces. PHB film and powder were completely degraded after 6 and 3 d cultivation, respectively at 50 degrees C. Scanning micrographs showed adherence of the microbial cells to the entire film surface, indicating that biodegradation occurs by colonization of the PHB surface. The film was degraded both by microbial attack and by the action of an extracellular enzyme secreted by the microorganism. The strain can also degrade poly(ethylene succinate), poly(ester carbonate), polycaprolactone and poly(butylene succinate), but to a lesser extent.

Biodegradation, Environmental↗
PubMed1999

Metabolic engineering of Arabidopsis and Brassica for poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer production.

Poly(hydroxyalkanoates) are natural polymers with thermoplastic properties. One polymer of this class with commercial applicability, poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) can be produced by bacterial fermentation, but the process is not economically competitive with polymer production from petrochemicals. Poly(hydroxyalkanoate) production in green plants promises much lower costs, but producing copolymer with the appropriate monomer composition is problematic. In this study, we have engineered Arabidopsis and Brassica to produce PHBV in leaves and seeds, respectively, by redirecting the metabolic flow of intermediates from fatty acid and amino acid biosynthesis. We present a pathway for the biosynthesis of PHBV in plant plastids, and also report copolymer production, metabolic intermediate analyses, and pathway dynamics.

Acyl Coenzyme A↗
PubMed2006

Genome sequence of the bioplastic-producing "Knallgas" bacterium Ralstonia eutropha H16.

The H(2)-oxidizing lithoautotrophic bacterium Ralstonia eutropha H16 is a metabolically versatile organism capable of subsisting, in the absence of organic growth substrates, on H(2) and CO(2) as its sole sources of energy and carbon. R. eutropha H16 first attracted biotechnological interest nearly 50 years ago with the realization that the organism's ability to produce and store large amounts of poly[R-(-)-3-hydroxybutyrate] and other polyesters could be harnessed to make biodegradable plastics. Here we report the complete genome sequence of the two chromosomes of R. eutropha H16. Together, chromosome 1 (4,052,032 base pairs (bp)) and chromosome 2 (2,912,490 bp) encode 6,116 putative genes. Analysis of the genome sequence offers the genetic basis for exploiting the biotechnological potential of this organism and provides insights into its remarkable metabolic versatility.

Aerobiosis↗
PubMed2001

Kinetic analysis on formation of poly(3-hydroxybutyrate) from acetic acid by Ralstonia eutropha under chemically defined conditions.

Batch cultures of Ralstonia eutropha in chemically defined media with acetic acid (HAc) as the sole carbon source were conducted to investigate acetate utilization, formation of poly(3-hydroxybutyrate) (PHB) and growth of active biomass (ABM) under different carbon to nitrogen (C/N) weight ratios. The specific acetate utilization rate based on ABM approached 0.16 g/g ABM h(-1), which was not affected very much by the extracellular HAc concentration from 1 to 5 g/l, but was affected by the C/N weight ratio. A low C/N ratio or high nitrogen supply sped up the specific acetate utilization rate to produce more ABM and less PHB. A high HAc concentration (>6 g/l), however, depressed acetate utilization as well as the ABM growth and PHB formation. A high cell mass concentration enhanced the tolerance of R. eutropha to the toxicity of HAc at pH 7 to 8.5. The viscosity-average molecular size of PHB generally increased first and then declined in batch cultures. Larger PHB molecules and less PHB per ABM were produced at a low C/N ratio with enough nutrient nitrogen than those under a high C/N ratio with less nutrient nitrogen available.

Acetic Acid↗