Studies on copolyester synthesis by Rhodococcus ruber and factors influencing the molecular mass of polyhydroxybutyrate accumulated by Methylobacterium extorquens and Alcaligenes eutrophus (original) (raw)
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The bacterial copolyester poly-(3-hydroxybutyrate-co-3-hydroxyvalerate) was produced with Alcaligenes eutrophus DSM 545 from glucose and sodium propionate in a fed-batch fermentation with both nitrogen limitation and low dissolved-oxygen concentrations. When the dissolved-oxygen content was kept between 1 and 4% of air saturation during the polymer accumulation phase, the yield of 3-hydroxybutyrate (3HB) monomer from glucose was not affected, but the propionate-to-3-hydroxyvalerate (3HV) monomer yield was two to three times (0.48 to 0.73 mol of 3HV mol of propionate consumed ؊1 ) that observed in a control experiment (0.25 mol mol ؊1 ), where the accumulation-phase dissolved-oxygen concentration was 50 to 70% of air saturation. The overall polymer productivity of the fermentation was somewhat decreased by low dissolved-oxygen contents, owing to a slower 3HB production rate. The effect of a low dissolved-oxygen concentration is probably attributable to a reduction of the oxygen-requiring decarbonylation of propionyl-coenzyme A (CoA) to acetyl-CoA.
Macromolecules, 2000
Two novel bacterial poly(3-hydroxyalkanoates) (PHAs) with either 3-hydroxy-7-oxooctanoate (HOO) and 3-hydroxy-5-oxohexanoate (HOH) or 8-acetoxy-3-hydroxyoctanoate (AHO), 6-acetoxy-3hydroxyhexanoate (AHH), and 4-acetoxy-3-hydroxybutyrate (AHB) monomer units were produced at pilot scale. For the biosynthesis of these PHAs Pseudomonas oleovorans was cultivated at a 24 L scale in two-liquid-phase fed-batch processes using mineral salts medium and mixtures of 2-octanone/octane or n-octylacetate/octane as carbon sources. The bacterial accumulation of the polyesters was induced by nitrogen starvation and the addition of the substrate mixtures. Under these conditions, 26 and 45 g of PHA were isolated. The PHAs contained 10.3 and 3.3 mol % of oxo and acetoxy group monomers. Physical characterization was done with respect to their molecular weights, and thermal properties and similar results were found as for octane-based PHA. All monomer units have been identified by 2D 1 H and 13 C heteronuclear correlated NMR spectroscopy (HSQC, HMBC, and HSQC-TOCSY), and the composition of the copolyesters was quantified from 1 H NMR spectra.
World Journal of Microbiology Biotechnology, 2005
Production of poly(3-hydroxybutyric acid) [P(3HB)] by Rhodopseudomonas palustris SP5212 isolated in this laboratory has been optimized under phototrophic microaerophilic conditions. Cells grown in malate medium accumulated 7.7% (w/w) P(3HB) of cellular dry weight at the early stationary phase of growth. The accumulated P(3HB) however, attained 15% (w/w) of cellular dry weight when acetate (1.0%, w/v) was used as the sole carbon source under nitrogen-limiting conditions. Synthesis and accumulation of polymer was favoured by sulphate-free conditions and at a phosphate concentration sub-optimal for growth. The polymer content of cells was increased drastically (34% of cellular dry weight) when the acetate containing medium was supplemented with n-alkanoic acids. Compositional analysis by H 1 NMR revealed that these accumulated polymers were composed of 3-hydroxybutyric acid and 3-hydroxyvaleric acid (3HV). The contents of 3HV in these copolymers ranged from 14 to 38 mol%. Materials and methods Bacterial strain and cultural conditions Rhodopseudomonas palustris SP5212 isolated from a sludge sample collected from a pond of the Hooghly
Frontiers in Bioengineering and Biotechnology
Microbial polyhydroxyalkanoates (PHA) are biodegradable and biocompatible bio-based polyesters, which are used in various applications including packaging, medical and coating materials. In this study, an extremophilic hydrocarbonoclastic bacterium, previously isolated from saline sediment in the Tunisian desert, has been investigated for PHA production. The accumulation of intracellular PHA granules in Halomonas desertis G11 was detected by Nile blue A staining of the colonies. To achieve maximum PHA yield by the strain G11, the culture conditions were optimized through response surface methodology (RSM) employing a Box-Behnken Design (BBD) with three independent variables, namely, substrate concentration (1–5%), inoculum size (1–5%) and incubation time (5–15 days). Under optimized conditions, G11 strain produced 1.5 g/L (68% of DCW) of PHA using glycerol as a substrate. Application of NMR (1H and 13C) and FTIR spectroscopies showed that H. desertis accumulated PHA is a poly-3-hydr...
Biotechnology and Bioengineering, 2001
A recombinant strain of Salmonella enterica serovar Typhimurium (mutant in propionate-activation activity) was metabolically engineered to control the composition of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), a polyhydroxyalkanoate copolymer with commercially desirable properties. A gene (prpE) encoding propionyl-CoA synthetase was placed under the control of the IPTG-inducible taclacUV5 promoter (P taclacUV5 ) while the polyhydroxyalkanoate synthesis operon (phaBCA) from Acinetobacter sp. RA3849 was coexpressed under the control of the arabinose-inducible araBAD promoter (P BAD ). S. enterica, harboring both constructs, was grown in medium containing a fixed substrate concentration and the composition of the copolymer was varied between 2 mol% and 25 mol% 3-hydroxyvalerate by controlling the IPTG level in the medium. This "dial-a-composition" system should find application in cases where the substrate concentration of a feedstream for PHBV bioplastic production is not adjustable.
Scientific reports, 2016
While poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] is a biodegradable commodity plastic with broad applications, its microbial synthesis is hindered by high production costs primarily associated with the supplementation of related carbon substrates (e.g. propionate or valerate). Here we report construction of engineered Escherichia coli strains for direct synthesis of P(3HB-co-3HV) from an unrelated carbon source (e.g. glucose or glycerol). First, an E. coli strain with an activated sleeping beauty mutase (Sbm) operon was used to generate propionyl-CoA as a precursor. Next, two acetyl-CoA moieties or acetyl-CoA and propionyl-CoA were condensed to form acetoacetyl-CoA and 3-ketovaleryl-CoA, respectively, by functional expression of β-ketothiolases from Cupriavidus necator (i.e. PhaA and BktB). The resulting thioester intermediates were channeled into the polyhydroxyalkanoate (PHA) biosynthetic pathway through functional expression of acetoacetyl-CoA reductase (PhaB) f...
Journal of Biotechnology, 2007
One of the main limitations in bacterial polyhydroxyalkanoate (PHA) production with mixed cultures is the fact that primarily polyhydroxybutyrate (PHB) homopolymers are generated from acetate as the main carbon source, which is brittle and quite fragile. The incorporation of different 3-hydroxyalkanoate (HA) components into the polymers requires the addition of additional carbon sources, leading to extra costs and complexity. In this study, the production of poly(3-hydroxybutyrate (3HB)-co-3-hydroxyvalerate (3HV)-co-3-hydroxy-2-methylvalerate (3HMV)), with 7-35 C-mol% of 3HV fractions from acetate as the only carbon source was achieved with the use of glycogen accumulating organisms (GAOs). An enriched GAO culture was obtained in a lab-scale reactor operated under alternating anaerobic and aerobic conditions with acetate fed at the beginning of the anaerobic period. The production of PHAs utilizing the enriched GAO culture was investigated under both aerobic and anaerobic conditions. A polymer content of 14-41% of dry cell weight was obtained. The PHA product accumulated by GAOs under anaerobic conditions contained a relatively constant proportion of non-3HB monomers (30 ± 5 C-mol%), irrespective of the amount of acetate assimilated. In contrast, under aerobic conditions, GAOs only produced 3HB monomers from acetate causing a gradually decreasing 3HV fraction during this aerobic feeding period. The PHAs were characterized by gel permeation chromatography (GPC) and differential scanning calorimetry (DSC). The data demonstrated that the copolymers possessed similar characteristics to those of commercially available poly(3HB-co-3HV) (PHBV) products. The PHAs produced under solely anaerobic conditions possessed lower melting points and crystallinity, higher molecular weights, and narrower molecular-weight distributions, compared to the aerobically produced polymers. This paper hence demonstrates the significant potential of GAOs to produce high quality polymers from a simple and cheap carbon source, contributing considerably to the growing research body on bacterial PHA production by mixed cultures.
2007
Kemavongse, K., Prasertsan, P., Upaichit, A. and Methacanon, P. Effect of co-substrate on production of poly-β-hydroxybutyrate (PHB) and copolymer PHBV from newly identified mutant Rhodobacter sphaeroides U7 cultivated under aerobic-dark condition Songklanakarin J. Sci. Technol., 2007, 29(4) : 1101-1113 Photosynthetic bacterial mutant strain U7 was identified using both classical and molecular (16S rDNA) techniques to be Rhodobacter sphaeroides. The glutamate-acetate (GA) medium containing sodium acetate and sodium glutamate as carbon and nitrogen sources was used for production of poly-β-hydroxybutyrate (PHB) from R. sphaeroides U7 cultivated under aerobic-dark condition (200 rpm) at 37oC. Effect of auxiliary carbon sources (propionate and valerate) and concentrations (molar ratio of 40/0, 40/20, 40/40 and ORIGINAL ARTICLE 1M.Sc. (Biotechnology), 2Ph.D. (Biotechnology), Assoc. Prof., 3Dr. Techn (Technical Chemistry), Department of Industrial Biotechnology, Faculty of Agro-Industry,...