• KSBB Journal
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• 제15권6호
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• pp.556-559
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• 2000

Cyclodextrin Glucanotransferase(EC 2.4.1.19 : 1,4-${\alpha}$-glucano) transferase, cyclizing; CGTase) can be separated and recovered in an aqueous two-phase system composed of poly(ethylene glycol)(PEG)/dextran and PEG/salt. In an aqueous two-phase system consisting of PEG 35000 (5%) and dextran T2000 (7%), all cell and debris were collected at the interphase. CGTase partitioned to the denser dextran phase at an yield of 83.4%. On the other hand, in an aqueous two-phase system consisting of PEG 35000 (10%) and sodium phosphate (15%), CGTase partitioned to the denser salt phase at an yield of 95.5%. In order to recover CGTase using an aqueous two-phase system, the PEG/salt system proved to be more efficient than the PEG/dextran system in terms of yield and cost.

• 한국미생물·생명공학회지
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• 제20권3호
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• pp.344-347
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• 1992

Bacillus stearothermophilus No.239의 돌연변이주 MNNG 8이 생산하는 CGTase를 사용하여 감자전분을 동시 액화, cyclodextrin(CD) 생산을 하였다. 고농도(30)의 감자전분이 29의 수율로 CD로 전환 되었으며 그 때의 조건은 pH 6.0, $80^{\circ}C$, 4.3mM, $CaCl_2$, $40^{\circ}C$에서 1g의 전분당 3.0 DAU의 CGTase를 첨가하는 것이다.

• 한국미생물·생명공학회지
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• 제30권4호
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• pp.293-297
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• 2002

효모 S. cerevisiae에서 B. stearothermophilus 유래의 CGTase를 발현 생산하였으며, 분비, 생산된 단백질을 정제하여 그 특성을 조사하였다. 재조합 효모 S. cerevisiae 2805/pVT- CGTS가 생산하는 CGTase의 분자량은 효모에서 발현될 때 고당쇄가 부가되어 야생형의 68kDa에 비해 15-160% 증가된 약 78-178 kDa으로 나타났다. 효모 S. cerevisiae에서 발현된 CGTase의 효소반응 최적활성조건은 pH7.0, $65^{\circ}C$였고, 열안정성에 있어서 $75^{\circ}C$에서 약 90%의 잔존활성을 가질 정도로 내열성이 개선되었다. 효모 S. cerevisiae에서 발현된 CGTase는 5% soluble starch를 기질로 약 40.2%의 CD 전환율 및 3 : 6 : 1의 $\alpha$-, $\beta$-, ${\gamma}$-CD의 생산 비율을 나타내어 야생형과 별다른 변화가 없었다.

• 한국미생물·생명공학회지
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• 제19권2호
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• pp.163-170
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• 1991

Production of cyclodextrin (CD) directly from raw corn starch without liquefaction using cyclodextrin glucanotransferase (CGTase) was carried out in an agitated bead reaction system. Similar CD yield and production rate comparable with those of conventional method using liquefied starch were obtained. Especially high purity-CD in the reaction mixture without accumulation of malto-oligosaccharides was obtained. The maximum 54g/l of CD was obtained at raw starch concentration of 200g/l. CD yield was inversely proportional to raw starch concentration, and conversion yield was 0.48 at substrate concentration of 100g/l. The optimal amount of enzyme (CGTase unit/g raw starch) was found to be around 6.0. Granular structure of raw starch degraded by CGTase was observed by SEM in order to investigate the enhancing mechanism, along with those of acid or alkali pretreated raw starch, amylose, and amylopectin. Kinetic constants of CGTase on raw starch in an agitated bead reaction system were evaluated, and CGTase was competitively inhibited by CD.

• 한국식품영양학회지
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• 제8권3호
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• pp.253-261
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• 1995

A strain of alkalophilic Bacillus sp. C-21 has been Isolated from sold. The strain was capable of producing large amount of cyclodextrin glycosyltransferase (CGTase) in the high alkaline pH medium. The preferable medium composition was determined to be as follows : 1.0% soluble starch, 1.0% peptone, 0.5% yeast extract, 0.1% K2HP04, 0.02% MgSO4.7H2O and 1.0% Na2CO3(pH 10.0) The highest enzyme production was observed after 30hours of cultivation at 33$^{\circ}C$. The optimum temperature and pH for the activity of crude enzyme were 6$0^{\circ}C$ and 6.0, respectively. The enzyme was stable between pH 6.0 and 9.6, and up to 55$^{\circ}C$.

• 한국식품영양과학회지
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• 제27권4호
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• pp.609-617
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• 1998

A microorganism capable of producing high level of extracellular cyclodextrin glucanotransferase(EC 2.4.1.19 ; CGTase) was isolated from Kimchi. 2-O-$\alpha$-D-glucopyranosyl L-ascorbic acid(AA-2G) was synthesized by transglycosylation reaction of CGTase using starch as a donor and L-ascorbic acid as an acceptor. The isolated strain S-6 was identified as Bacillus sp. S-6. The maximal CGTase production was observed in a medium containing 0.5% soluble starch, 1% yeast extract, 1% NaCO3, 0.1% K2HPO4, and 0.02% MgSO4 with initial pH 8.0. The strain was cultured at 37$^{\circ}C$ for 40 hr with reciprocal shaking. Using the culture supernatant as crude enzyme, the optimal pH and temperature of the CGTase activity of this strain were 7.0 and 4$0^{\circ}C$. In the effects of pH and temperature on the stability of the enzyme, the enzyme was stable in the range of pH 6.0~10.0 and up to 45$^{\circ}C$, respectively.

• 한국식품영양과학회지
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• 제29권1호
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• pp.49-56
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• 2000

The 2-O-$\alpha$-D-glucopyranosyl L-ascorbic acid (AA-2G) which was enzymatically glucosylated with the cyclodextrin glucanotransferase (CGTase) [EC 2.4.1.19] from Bacillus sp. JK-43 has been reported previously. The presnet experiments examined the optimal conditons for the productio of AA-2G from AA and soluble starch, and characterized the properties of the CGTase from Bacillus sp. JK-43. The reaction mixture for the maximal production of AA-2G was followings; 12% total substrate concentration, 1,400 usits/mL of CGTase and a mixing ratio of 2 : 3(g or AA : g of soluble starch). Under this condition, 1.76mM of AA-2G, which corresponded to 2.53% yield based on AA, was produced after incubation for 24hrs at 45$^{\circ}C$ (pH 5.5). The optimum pH and temperature for the CGTase activity were 6.0 and 45$^{\circ}C$, respectively. The enzyme was stable at pH 5.5 to 9.5, and at temperature up to 5$0^{\circ}C$. The thermostability of the enzyme could be enhanced up to 6$0^{\circ}C$ by the addition of 30mM CaCl2.

• 한국미생물·생명공학회지
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• 제17권4호
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• pp.370-378
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• 1989

토양으로부터 역가높은 CGTase를 분비하는 호알칼리성 미생물을 분리하였으며, 동정 결과 Bacillus circulans로 판정되었다. 배양액 중의 CGTase를 ammonium sulfate 침전, DEAE-Sephadex 그리고 Sephadex G-100 column chromatography로 분리, 정제하여 단일 단백질 band를 얻었다. 정제된 CGTase의 분자량은 약 93,000, 최적 pH와 온도는 6.0, $50^{\circ}C$였으며, pH와 온도안정성은 5.5-11, $65^{\circ}C$까지였다. Soluble starch를 기질로 할 때의 $V_{max}$ 와 $K_{m}$ 값은 각각 0.16$\mu$mole $\beta$-CD/min, 14.3mg soluble starch/mi이였고 24시간 반응액의 $\alpha$-：$\beta$-:${\gamma}$-CD 의 생성비율은 1：8.1：1.9로서 $\beta$-CD를 우선적으로 합성하였다. 기질로 glucose와 maltose를 사용하였을 때 CD합성작용이 없었으며, sweet potato 그리고 cornstarch를 사용하였을 때 가장 높은 CD합성작용을 보였다. 어느 수준 이상의 과다한 CGTase 첨가경우에는 $\alpha$-CD생성이 급격히 증가하였다. 또한 정제된 CGTase는 stevioside에로의 당전이성을 갖고 있었다.

• 한국식품과학회지
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• 제29권5호
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• pp.969-973
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• 1997

고정화 CGTase를 이용하여 당전이 스테비오사이드를 연속적으로 제조하기 위하여 몇가지 생물반응기의 적용가능성을 조사하였다. 연속형 반응기로는 packed-bed reactor(PBR)가 적합하였다. 이때 specific productivity는 0.4 g/hr-g immobilized enzyme으로 회분식반응기의 50% 정도이나 연속생산이 가능한 장점이 있었다. PBR의 운전조건이고 $10{\sim}60\;cm/hr$의 선속도 범위내에서 스테비오사이드 전환률 70% 이상을 보이는 기질의 유속은 공간속도로 $0.5{\sim}0.7\;hr^{-1}$이었다. 고정화 효소의 운전 안정성은 $47^{\circ}C$에서 30%의 기질로 2주 일간 연속 사용이 가능하고, 반응기의 control방법으로 반응기 상부에서 20%의 효소를 교체하면 20일 이상 전환률이 유지되었다.

• Journal of Microbiology and Biotechnology
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• 제1권1호
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• pp.63-69
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• 1991

Performance of column type bioreactor packed with immobilized cyclodextrin glucanotransferase (CGTase) on chitosan and Amberite IRA 900 was evaluated for cyclodextrin(CD) production. For CGTase immobilized on chitosan, the maximum CD conversion yield of 42% was achieved at the range of 88-168 units of immobilizied CGTase per gram of chitosan, retention time of 0.3 hr, and from 5.0% (w/v) of partially cyclized soluble starch. On the other hand, for CGTase immobilized on Amberite IRA 900, the maximum conversion yield of 40% was obtained at the range of 3.6-11.0 units of immobilized CGTase per gram of carrier and retention time of 1.2 hr from 5.0% of substrate. Above CD conversion yields are almost identical level with that can be obtained with soluble CGTase of 47%. The productivities of bioreactor packed with immobilized CGTase were 17.0g of CD/lㆍhr for amberite IRA 900 and 15.5g of CD/lㆍhr for chitosan. The partially cyclized starch with soluble CGTase were more suitable as substrate to achieve better CD conversion yield, and 5% (w/v) of partially cyclized soluble starch containing 10% (w/w) of CD was found to be most suitable to obtain maximum CD conversion yield.