• Microbiology and Biotechnology Letters
• /
• v.21 no.3
• /
• pp.221-228
• /
• 1993

In order to achieve poly-beta-hydroxybutyric acid (PHB) production using recombinant DNA in various host bacterial cells, the isolation of genes for PHB biosynthesis was attempted. As a result, a 5.2kb DNA fragment containing phbCAB operon of Alcaligenes eutrophus was isolated by colony hybridization using synthetic oligodeoxyribonucleotides as probes. The constructed recmbinant plasmid pSK(+)-phbCAB operon was transferred to Escherichia coli, and the obtained transformant accumulated considerable amount of PHB.

• Korean Chemical Engineering Research
• /
• v.54 no.5
• /
• pp.719-721
• /
• 2016

Formate has been considered as an environmentally sustainable feedstock that can be used to accelerate the production of valuable chemicals. This study presents brief results of the formatotrophic production of Poly-${\beta}$-hydroxybutyric acid (PHB) by Methylobacterium sp. To evaluate the production of PHB, five species of Methylobacteria were tested using formate as the sole carbon and energy source. Methylobacterium chloromethanicum CM4 exhibited the highest productivity of PHB, which showed 1.72 g/L PHB production, 32.4% PHB content, and 0.027 g-PHB/g-formate PHB yield. These results could be used for the formatotrophic production of PHB with the concurrent reduction of $CO_2$ to formate.

• Applied Biological Chemistry
• /
• v.34 no.3
• /
• pp.273-278
• /
• 1991

Methanol assimilating bacterium, Pseudomonas sp. ILS-003 was used to investigate the optimum conditions for the production of $poly-{\beta}-hydroxybutyric$ acid from methanol. For PHB production, the optimum initial pH was 6.4 and the optimum temperature was $30^{\circ}C$. Also the optimum methanol concentration was found to be 1.0%(v/v). In the PHB production, $(NH_4)_2SO_4$ was the most effective nitrogen source and the optimum concentration was 0.8 g/l, which was eqivalent to 17.4 in C/N ratio. Also, deficiency of the 2 valence metal ions in the medium had stimulating effect on PHB accumulation. Under the optimum substrate concentration, successive feeding of 0.25%(v/v) methanol was the most effective on PHB production. Under the optimum conditions, 1.94 g/l of PHB and 2.78 g/l of dry biomass were produced in 96 hours, and the yield was 69.8%(w/w).

• Microbiology and Biotechnology Letters
• /
• v.20 no.4
• /
• pp.363-370
• /
• 1992

Microorganisms capable of accumulating poly-p-hydroxybutyric acid(PHB) were isolated from soil by enrichment culture technique. Among them, the strain designated as FL-027 had high PHB productivity and was identified as Alcaligenes. The optimal medium composition for cell growth was 8.0 $g/\ell$ of fructose and 3.0 $g/\ell$ of $(NH_4)_2S0_4$, equivalent to C/N ratio 5.04 at pH 7.0 and $30^{\circ}C$. To investigate the optimal conditions for the PHB accumulation, we divided the process into two stages; the first stage for the growth of the cell in nutrient-rich medium and the second stage for the PHB accumulation in nutrient-deficiency medium. The optimal conditions for PHB accumulation were 8.0 $g/\ell$ of fructose and 0.25 $g/\ell$ of $(NH_4)_2S0_4$, equivalent to C/N ratio 60 at pH 6.5 and $30^{\circ}C$. PHB accumulation was stimulated by deficiency of nutrients such as $NH_4^+$, $Ca^{2+}$, $SO_4^{2+}$ in medium. Among them. $NH_4^+$ deficiency was chosen because of its effectiveness. We found the inhibition of cell growth by fructose in batch culture. In order to keep the fructose concentration at an optimal level, intermittent feeding fed-batch culture was employed, and the cell concentration as high as 10.83 $g/\ell$ whose PHB content was responsible for 43% of the dry cell weight. The purified PHB was identified as homopolymer of 3-hydroxybutyric acid by using IR and $^1H-NMR$.

• KSBB Journal
• /
• v.6 no.1
• /
• pp.79-83
• /
• 1991

Using PHB biopolymer as polymer matrix, the release mechanism of a model drug, silver sulfadiazine was studied. The release behavior actually conformed to the Higuchi's diffusion controlled model. The release rate was delayed with an increasing proportion of PHB, whereas decreased as glycerine concentration incresed. The release rate was increased as the polymer matrix thickness increased.

• KSBB Journal
• /
• v.11 no.2
• /
• pp.246-253
• /
• 1996

An obligatory methanotroph Methylosinus trichosporium OB3b was cultivated for the production of poly-${\beta}$-hydroxybutyric acid(PHB) in shake-flask using liquefied natural gas(LNG) as the sole source of carbon and energy. The maximal specific growth rate decreased by 40% using LNG compared with that obtained with pure methane. This is attributed to the inhibition by ethane and propane presents in the LNG as impurities. For the production of PHB, two-stage culture separating the production stage from the growth stage was carried out. PHB accumulation was observed after switching nutrient-sufficient to nutrient-limited condition of non-carboneous component (NO3-, PO43-, K+, Na+, Fe2+, or Mg2+). The limitation of K+ or Mg2+ resulted in relatively high PHB content, but the highest content was obtained by nitrate limitation. The optimal pH and temperature for PHB accumulation was 7.0 and $30^{\circ}C$. Under the optimal condition the maximal PHB content was about 45% after 4-day cultivation.

• Microbiology and Biotechnology Letters
• /
• v.18 no.3
• /
• pp.273-279
• /
• 1990

For poly- $\beta$ -hydroxybutyrate (PHB) production, a pink-pigmented facultative methylotrophic bacterium (PPFM) P-10 was newly isolated from soils through methanol-enrichment culture. The optimal medium composition for cell growth was 1.0% (vlv) of methanol as carbon source and l.Og/l of ,TEX>$NH_4Cl$, equivalent to C/N ratio of 13.2 at pH 7.0 and $30^{\circ}C$. To investigate the optimal condition for YHB accumulation, two-stage culture technique was adopted; first stage for cell growth and second stage for accumulation of PHB providing unbalanced growth conditions. The optimal PHB accumulation was 1.0% (vIv) of methanol and 0.26gll of $NH_4Cl$, C/N of 50.8 at pH 6.0. To overcome methanol inhibition on cell growth, intermittent feeding fed-batch culture technique was employed, and the cell concentration as high as 14gll with 40% of PHB was achieved. The purified PHB was identified using IR and $1^H NMR$ as homopolymer of 8hydroxybutyric acid. The absorption spectrum of extracted pink colored microbial pigment was alsa investigated.

• Journal of Microbiology and Biotechnology
• /
• v.7 no.4
• /
• pp.229-236
• /
• 1997

A gene, $phbC_{2.4.1}$ encoding poly-3-hydroxybutyric acid (PHB) synthase of Rhodobacter sphaeroides 2.4.1 was cloned by employing heterologous expression in Escherichia coli. R. sphaeroides chromosomal DNA partially digested with MboI was cloned in pUC19 followed by mobilization into E. coli harbouring $phbA,B_{AC}$ in pRK415, which code for ${\beta}$-ketothiolase and acetoacetyl CoA reductase of Alcaligenes eutrophus, respectively. Two E. coli clones carrying R. sphaeroides chromosomal fragment of $phbC_{2.4.1}$ in pUC19 were selected from ca. 10,000 colonies. The PHB-producing colonies had an opaque white appearance due to the intracellular accumulation of PHB. The structure of PHB produced by the recombinant E. coli as well as from R. sphaeroides 2.4.1 was confirmed by [$H^{+}$]-nuclear magnetic resonance (NMR) spectroscopy. Restriction analysis of the two pUC19 clones revealed that one insert DNA fragment is contained as a part of the other cloned fragment. An open reading frame of 601 amino acids of $phbC_{2.4.1}$ with approximate M.W. of 66 kDa was found from nucleotide sequence determination of the 2.8-kb SaiI-PstI restriction endonuclease fragment which had been narrowed down to support PHB synthesis through heterologous expression in the E. coli harbouring $phbA,B_{AC}$. The promoter (s) of the $phbC_{2.4.1}$ were localized within a 340-bp DNA region upstream of the $phbC_{2.4.1}$ start codon according to heterologous expression analysis.

• Korean Journal of Microbiology
• /
• v.20 no.4
• /
• pp.195-200
• /
• 1982

Rhizobium japonicum was isolated from the nodule of soybean root grown at the reclaimed tidal land in Kang-Wha island. The effect of pH and salt concentration to the viability of the isolated strain were examined in relationship between microbial growth and accumulation of PHB. Optimal pH value for the good viability of the isolated strain was 7.0 and also, at 5.0 and 6.0 viability was favorable to large extent, but 9.0 was unfavorable. Examined the effect of salt concentration treated two times as of the salinity in the reclaimed tidal land, viability of the isolated strain showed about 30 to 40%. And also in treatment with NaCl(40g/l) whatever the pH value adopted, viability was mostly less than 10%. The amount of accumulated PHB was relatively high at low pH value(5-6) and at high salt concentrration, respectively.

• Journal of Life Science
• /
• v.18 no.12
• /
• pp.1625-1630
• /
• 2008

A microorganism capable of producing high level of poly-3-hydoxybutyrate (PHB) from xylose was isolated from soil. The isolated strain J-65 was identified as Bacillus megaterium based on the morphological, biochemical and molecular biological characteristics. The optimum temperature and pH for the growth of B. megaterium J-65 were $37^{\circ}C$ and 8.0, respectively. The optimum medium composition for the cell growth was 2% xylose, 0.25% $(NH_4)_2SO_4$, 0.3% $Na_2HPO_4{\cdot}12H_2O$, and 0.1% $KH_2PO_4$. The optimum condition for PHB accumulation was same to the optimum condition for cell growth. Copolymer of ${\beta}$-hydroxybutyric and ${\beta}$-hydroxyvaleric acid was produced when propionic acid was added to shake flasks containing 20 g/l of xylose. Fermenter culture was carried out to produce the high concentration of PHB. In batch culture, cell mass was 9.82 g/l and PHB content was 35% of dry cell weight. PHB produced by B. megaterium J-65 was identified as homopolymer of 3-hydoxybutyric acid by GC and NMR.