• KSBB Journal
• /
• v.9 no.4
• /
• pp.365-371
• /
• 1994

The effects of ratios of initial concentration of carbon source to the initial concentration of nitrogen source in the fermentation media on both the yields of PHB fermentation variables and the accumulation of poly-${\beta}$-hydroxybutyric acid(PHB) were investigated. The fermentation media were composed of the combination of varing glucose concentrations, 10, 20, 25, 30, 40, $50g/\ell$ and the NH4Cl concentrations 0.33, 0.4, 0.5, 1.5, 3, $5g/\ell$. The yield of biomass on glucos, Yx/s, decreased very slowly according to the increase of the ratio of C to N. And the yield became constant at 0.35(g biomass/g glucose) with the ratio higher than 70. The yield of residual biomass, Yx/s, also decreased with the ratio of C to N and finally showed a constant value of 0.065(g residual biomass/g glucose) when the ratio was higher than 65. In accordance with the augmentation of the ratio, the yield of PHB, YPHB/S, however, increased and showed the maximum value of 0.35 (g PHB/g glucose) between 40 and 60 of the ratio. The maximum yield of PHB to the change of biomass, YPHB/S, was 0.87(g PHB/g biomass), and the yie1d YPHB/RX, was 4.2(g PHB/g residual biomass). The maximum accumulation percent of PHB to the final biomass was 81% when the ratio was higher than 67.

• Journal of Microbiology and Biotechnology
• /
• v.11 no.2
• /
• pp.310-316
• /
• 2001

Poly-3-hydroxybutyrate (PHB) synthase catalyzed the last enzymic step to synthesize the intracellular PHB of Rhodobacter sphaeroides. No PHB was detected when the phbC coding for PhB synthase was interrupted, and its expression was regulated at the level of transcription. The cellular PHB content increased about four- to six-fold during the growth transition from the exponential to the early stationary phase under both aerobic and photoheterotrophic conditions. The PHB content during the aerobic growth seemed to be determined by the PhB synthase activity. However, the PHB synthase activity of photoheterotrophically grown cells did not correlate with the PhB content, suggesting a photoheterotrophic regulation different from the aerobic control. Thus, the PHB content of R. sphaeroides was regulated at the transcription level only under aerobic conditions.

• Microbiology and Biotechnology Letters
• /
• v.22 no.6
• /
• pp.614-620
• /
• 1994

The effects of growth temperature and nutritional components on the synthesis of poly-3-hydroxybutyric acid, PHB, by filamentation-suppressed recombinant Escherichia coli XL1-Blue (pSYL107) were studied. After culturing XL1-Blue(pSYL107) for 48 hours in complex medium at 30$\circ$C, 7Al g/l of PHB could be obtained with the PHB content and PHB yield of 82% and 0.371 g PHB/g glucose, respectively. Lower concentration of PHB(3.2 g/l) was obtained when cultu- red at 37$\circ$C, which seemed to be due to the instability of this strain having amplified FtsZ activity. The PHB concentration of 3.75 g/l was obtained after culturing 60 hours in R medium supplemen- ted with 20 g/l glucose at 30$\circ$C, which was more than twice higher than that obtained with XL1-Blue(pSYL105). This suggested that the enhancement of PHB synthesis by suppressing filamenta- tion was more significant in a defined medium than complex medium. PHB synthesis could be further enhanced by supplementing a small amount of various complex nitrogen sources. When 5 g/l of beef extract was added to a defined medium, PHB concentration, PHB content, and PHB yield obtained after 60 hours of cultivation at 30$\circ$C were 7.46 g/l, 86%, and 0.375 g PHB/g glucose,respectively.

• KSBB Journal
• /
• v.10 no.2
• /
• pp.176-182
• /
• 1995

The environmental and physiological factors affecting the production of exopolysaccharide (Methylan) and Poly-${\beta}$-hydroxybutyrate(PHB) by Methylobacterium organophilum were investigated. The maximum PHB content was obtained at $38^{\circ}C$ whereas maximum polysaccharide concentration was $3.54g/\ell$ at $30^{\circ}C$. Optimum pH was pH 7-8 for PHB production and pH 6-7 for polysaccharide production, respectively. Under the condition of $Mo^{2+}, Mg^{2+} or Mn^{2+}$ limitation with nitrogenlimitation, the PHB accumulation was increased, whereas the polysaccharide production was decreased as compared with that of solenitrogenlimitation. Under the condition of sole K+ limitation, cell growth was significantly inhibited and no polysaccharide was produced. However, the PHB content was as high as 60% of dry cell weight. Effect of C/N ratios (methanol/ammonium) in the feeding solution was examined for the simultaneous production of polysaccharide and PHB. The higher ratio of C/N showed the lower cell growth, higher content of PHB in cells, and higher yield of polysaccharide.

• 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.

• 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.13 no.3
• /
• pp.477-481
• /
• 2003

The community escape response of Rhodobacter sphaeroides is exerted through the action of CerR and CerI, which code for a LuxR-type regulatory protein and acylhomoserine lactone synthase, respectively. Deletion of chromosomal DNA including cerR and cerI (mutant RI) or insertional interruption of cert (mutant AP3) resulted in two-fold increase in the cellular poly-${\beta}$-hydroxybutyrate (PHB) content In comparison with the wild-type under aerobic growth conditions. The PHB synthase (PhbC) activities of the cer mutants were doubled, and the enzyme expression was regulated at the level of phbC transcription. Thus, CerR, possibly in response to autoinducer (AI), appears to modulate the PHB content of aerobically grown cells by downregulating phbC transcription.

• 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 $^1H-NMR$.

• Korean Journal of Environmental Biology
• /
• v.17 no.3
• /
• pp.249-255
• /
• 1999

The effect of surface roughness on biodegradability of poly (3-hydroxybutyrate) was investigated. The PHB film prepared by cooling the molten polymer slowly ($-0.5^{\circ}C$/min) had higher crystallinity and melting temperature than that prepared by quenching into liquid nitrogen followed by annealing at $90^{\circ}C$ for 2 hours. However, the former sample was found to degrade faster than the latter due to presence of microscopic crack. Roughening the surface of a PHB film by hot pressing under a coarse surfaced plate accelerated the bioerosion considerably of the sample in comparison with the sample having the same thermal history but smooth surface.

• Journal of Microbiology and Biotechnology
• /
• v.11 no.2
• /
• pp.333-336
• /
• 2001

An isolated phbC gene from Alcaligenes latus was reintroduced into the parent A. latus through the transformation process, and the effect of the amplified phbC gene on the biosynthesis of P(3-hydroxybutyrate-3-hydroxyvalerate) [P(3HB-3HV)] and P(3-hydroxybutyrate-4-hydroxybutyrate) [P(3HB-4HB)] in the transformant A. latus was investigated. The biosynthesis rate and content of the above copolymers increased up to 1.3-fold after enforcing its own phbC gene, and the molar fractions of 3HV and 4HB in P(3HB-3HV) and P(3HB-4HB) also changed remarkably from 35.0 to 48.0% and from 34.0 to 56.0%, respectively, showing a critical role of PHB synthase which catalyzes the polymerizing reactions between eiher 3HV or 4HB from precursor compounds and 3HB.