• Journal of Microbiology and Biotechnology
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• 제16권5호
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• pp.786-790
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• 2006

Cellular components of Lactococcus lactis ssp. lactis (heat-killed whole cells, cytoplasm, and cell walls) were tested for their in vivo immunopotentiating activity. Peritoneal macrophages from mice orally administered with heat-killed whole cells exhibited significantly greater phagocytic activity than the groups administered with cell-wall fraction or cytoplasm fraction. The cytotoxicity of natural-killer cells was the highest in the group administered with whole cells, and the production of cytokines ($IFN-\gamma$, IL-2, and IL-12) in spleen cells was significantly higher, when cellular components were injected, and it tended to be higher in the cell-wall and cytoplasm groups than in the whole-cell group. Interestingly, the cytokine production of Peyer's patch cells was high, when cytoplasm fractions were administered. These results demonstrate that whole cells and cytoplasm and cell-wall fractions of L. lactis ssp. lactis have immunopotentiating activities, which are related to the stimulation of Peyer's patches.

• Journal of Microbiology and Biotechnology
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• 제24권5호
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• pp.619-628
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• 2014

To identify lipase LipA (PFL_0617) from Pseudomonas protegens Pf-5, a lipA deletion mutant (Pf0617) and a complementary strain (Pf0617lipA) were constructed, and their effects on the lipase production were examined. Pf0617 remarkably decreased its whole-cell lipase activity, whereas Pf0617lipA made its whole-cell lipase activity not only restore to wild-type level but also get a further increment. However, the deletion and overexpression of lipA did not affect the extracellular lipase activity. In addition, the unbroken whole cells of these strains were able to catalyze the hydrolysis of membrane-permeable p-nitrophenyl esters, but could not hydrolyze the membrane-impermeable olive oil. These results confirmed that LipA was an intracellular lipase and Pf-5 could also be used as a natural whole-cell biocatalyst. To evaluate the potential of Pf-5 as a whole-cell biocatalyst and separately characterize the whole-cell LipA, the properties of the whole-cell lipases from Pf-5 and Top10lipA were characterized. The results demonstrated that both Pf-5 and Top10lipA exhibited high tolerance to alkaline condition, high temperature, heavy metal ions, surfactants, and organic solvents. Taken together, lipA can realize functional expression in E. coli Top10, and Pf-5 and Top10lipA as whole-cell biocatalysts may have enormous potential in applications.

• KSBB Journal
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• 제14권3호
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• pp.264-272
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• 1999

Mandelate pathway를 거치는 Pseudomonas putida(KCTC 1751)의 전세포 benzoylformate dcarboxylase를 이용하여 benzoylformate를 benzaldehyde로 변환하였고 성장배지의 조성이 cell내부에 축적되는 benzoylformate dcarboxylase의 양에 미치는 영향을 조사하였다. 전세포효소의 재사용을 위하여 calcumalginate 캡슐 고정회법을 이용하여 캡슐고정화 Pseudomonas putida를 제조하였다. 캡슐 고정화 미생물을 M3배지에서 3일간 배양한 후 M1배지에서 1일간 배양한 결과 77.75g/L의 미생물 건조중량을 얻었다. 캡슐 고정화 전세포 benzoyltormate decarboxylase의 비활생도는 자유배양에 의한 전세포효소의 비활성도에 비해 약 1/2값을 나타내었으며 캡슐 고정화 전세포 benzoyltormate decarboxylase를 20회 재사용시 20%의 실활을 보였으며 캡슐 재사용 30회 이후 미생물의 건조중량은 약 10% 감소를 보였다.

• 미생물학회지
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• 제24권3호
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• pp.276-287
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• 1986

ChIarella elliPsoidea를 rifampicin이 함유된 M4N 배지에 7일간 배양하였다. 배양기간 동안에 일정량 세포를 수확하여 생장율을 측정하였다. 이들 세포에서 분리된 엽록체로 부터 핵산과 RNA polymerase를 추출하여 함량을 염기별로, 분석, 활성도를 측정하여 핵산 합성에 비치는 rifampicin의 효과를 대조구와 비교하며 분석하였다. 생장 억제 효과를 나타내는rifampicin의 농도는 80ppm 이였다. Whole cell system과 분리한 엽록체에서의 DNA함량은 대조구에 비해 각각 70%, 60%의 감소를 나타내였다. Rifampicin은 RNA 염기 함양도 whole cell system에서는 46% 억제되었고 분리한 엽록체에서는 77% 저해효과가 관찰되었다. Rifampicin에 의한 RNA polymerase 활성도는 whole cell system에서는 10% 감소, 분리한 엽록체에서는 42% 억제를 나타내었다.

• Food Science and Biotechnology
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• 제14권3호
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• pp.405-409
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• 2005

Cellular components of Lactococcus lactis ssp. lactis (heat-killed whole cells, cytoplasm, and cell walls) were tested for their in vivo immunopotentiating activities. Peritoneal macrophages from mice injected intraperitoneally with cell-wall fractions exhibited significantly greater phagocytic activity than groups injected with whole cells or cytoplasm fraction. Cytotoxicity of natural-killer cells was highest in cytoplasm fractions. Production of cytokines (IFN-${\gamma}$, IL-2, IL-6, and IL-12) in spleen cells was significantly higher when cellular components were injected intraperitoneally, and tended to be higher in whole-cell and cytoplasm groups than in cell-wall group. These results demonstrate lactic acid bacteria whole cells and their cytoplasm and cell-wall tractions have immunopotentiating activities.

• Journal of Microbiology and Biotechnology
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• 제26권7호
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• pp.1206-1215
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• 2016

Ginsenosides are the major active ingredients in ginseng used for human therapeutic plant medicines. One of the most well-known probiotic bacteria among the various strains on the functional food market is Lactobacillus rhamnosus GG. Biocatalytic methods using probiotic enzymes for producing deglycosylated ginsenosides such as Rd have a growing significance in the functional food industry. The addition of 2% cellobiose (w/v) to glucose-free de Man-Rogosa-Sharpe broths notably induced β-glucosidase production from L. rhamnosus GG. Enzyme production and activity were optimized at a pH, temperature, and cellobiose concentration of 6.0, 40℃, and 2% (w/v), respectively. Under these controlled conditions, β-glucosidase production in L. rhamnosus GG was enhanced by 25-fold. Additionally, whole-cell homogenates showed the highest β-glucosidase activity when compared with disrupted cell suspensions; the cell disruption step significantly decreased the β-glucosidase activity. Based on the optimized enzyme conditions, whole-cell L. rhamnosus GG was successfully used to convert ginsenoside Rb1 into Rd.

• Archives of Pharmacal Research
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• 제3권1호
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• pp.7-12
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• 1980

Ampliciline was synthesized from 6-amino-pencillanic acid (6-APA) and D-.alpha. phenylglycine methyl ester by using amplicilin synthesizing enzyme from Peudomonas melanogenum (IAM 1655). The whole cell enzyme was immobilized by entrapping it in the polyacrylamide gel lattices. The polymer used in the enzyme entrapment was made from 150 mg per ml of acrylamide monomer and 8 mg per ml of N, N'-methylenebisacrylamide. About 200 mg/whole cell enzyme was mixed in the polymer for entrapment. The maximal activity retention after immobilization was 56%. The optimal pH values for the whole cell enzyme and the immobilized whole cell enzyme were 6.0 and 5.9, respectively. The optimal temperature for the enzyme activity were the same for both type of preparations. The enzyme stabilities against pH and heat increased for immobilized whole cell enzyme. Immobilized cell was more stable especially in the acidic condition while both type were found to be very suceptible to thermal inactivation at a temperature above 4.deg.C. The kinetic constants obtained from Lineweaver-Burk plot based on two substate reaction mechanism showed somewhat higher value for immobilized whole cell enzyme as compared to the whole cell enzyme : the Km value for 6-APA were 7.0 mM and 12.5 mM while Km values for phenylglycine methyl ester were 4.5 mM and 8.2 mM, respectively. Using the immobilized whole cell enzyme packed in a column reactor, the productivity of ampiciline was studied by varying the flow rate of substrate solution. At the space velocity, SV, 0.14 hr$^{-1}$ the conversion was 45%. Operational stability found in terms of half life was 30 hr at SV = 0.2 hr.

• KSBB Journal
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• 제11권4호
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• pp.398-404
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• 1996

전세포 효소 고정화 캡슐을 제조하기 위하여 Eschericia coli를 calcium alginate 캡슐내부에 접종하고 배양하였다. E. coli의 캡슐내부 건조중량이 1OOg/L에 달하였다. 생산배지에서 배양하는 동안 캡슐내부에 축적되는 미생물의 양이 많을수록 캡슐 내부에 축적되는 $\beta$galactosidase의 활성도 높았다. 생산배지에 금속이온 $Zn^{+2}를 2{\times}10^{-4} M $ 첨가함으로써 캡슐내부에 축척되는 ${\beta}$-galactosidase의 활성 을 25% 증가시킬 수 있었다. 캡슐제조시 해바라기유를 부피비로 2% 첨가함으로써 캡슐내부에 축적되 는 ${\beta}$-galactosidase의 활성을 10% 증가시킬 수 었었다. 부피산소전달계수, $k_La가 2.55h^- $ 안 플라스크 대신 $k_La가 82h^- $인 concentric air lift reactor 내에서 고정화 E. coli를 배양호농로써 캡슐내부의 전세포${\beta}$-galactosidase의 활성을 86% 증가시킬 수 있었다.

• KSBB Journal
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• 제26권3호
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• pp.199-205
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• 2011

Using tetrameric ${\beta}$-galactosidase as a model protein, anchoring motives were screened in Bacillus subtilis spore display system. Eleven spore coat proteins were selected considering their expression levels and the location in the spore coat layer. After chromosomal single-copy homologous integration in the amyE site of Bacillus subtilis chromosome, cotE and cotG were chosen as possible spore surface anchoring motives with their higher whole cell ${\beta}$-galactosidase activity. PAGE and Wester blot of extracted fraction of outer layer of purified spore, which express CotE-LacZ or CotG-LacZ fusion verified the existence of exact size of fusion protein and its location in outer coat layer of purified spore. ${\beta}$-galactosidase activity of spore with CotE-LacZ or CotG-LacZ fusion reached its highest value around 16~20 h of culture time in terms of whole cell and purified spore. After intensive spore purification with lysozyme treatment and renografin treatment, spore of BJH135, which expresses CotE-LacZ, retained only 1~2% of its whole cell ${\beta}$-galactosidase activity. Whereas spore of BJH136, which has cotG-lacZ cassette in the chromosome, retained 10~15% of its whole cell ${\beta}$-galactosidase activity, proving minor perturbation of CotG-LacZ, when incorporated in the spore coat layer of Bacillus subtilis compared to CotE-LacZ. Usage of Bacillus subtilis WB700, of which 7 proteases are knocked-out and thereby resulting in 99.7% decrease in protease activity of the host, did not prevent the proteolytic degradation of spore surface expressed CotG-LacZ fusion protein.

• Biotechnology and Bioprocess Engineering:BBE
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• 제6권1호
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• pp.67-71
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• 2001

Most of the Cyclodextrin glucanotransferase (Gtases) which have been produced from B. subtilis were found to be excreted from the cells during cultivation. Immobilized whole cell CGTase from B. subtilis was prepared by encapsulating the broth solution which had been concentrated ten times with a rotary vacuum evaporator. Cyclization activity of CGTase was reduced by about 10% during the concentrating process, however, its transglycosylation activity, to convert xylitol to glucosyl-xylitol, using dextrin as glucosyl donor, increased by a factor of 3 or 5.