• 한국미생물·생명공학회지
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• 제22권3호
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• pp.252-258
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• 1994

Transglycosylation of stevioside in the attrition coupled heterogeneous reaction system using raw starch as a glycosyl donor has significant advantages over conventional reaction systems using liquefied starch as a donor. The transglycosylation of stevioside under the presence of organic solvent showed that transglycosylation reaction occurs via two steps ; initially from raw starch to cyclodextrin(CD), and then followed by transglycosylation of produced CD. Comparison of the transglycosylation efficiency of c$\alpha $-, $\beta $, $\gamma $-CDs indicated that $\alpha $-, $\beta $-CD are mainly utilized as a glycosyl donor for following reaction. The reaction mechanism of transglycosylation between stevioside and CD proceeded according to random sequential bireactant mechanism. The equilibrium constant of transglycosylation reaction of cyclodextrin glucanotransferase wase also evaluated. The structure of transglycosylated stevioside was confirmed by TLC, and it was found that glycosyl group(G$_{1}, $ ~ G$_{4}$-glycosidic bond.

• 한국미생물·생명공학회지
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• 제21권5호
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• pp.461-467
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• 1993

Transglycosylation reaction of cyclodextrin glucanotransferase (CGTase) was analyzed in the attrition coupled heterogeneous reaction system using raw starch as a donor` and mono-, di-saccharide, and glycoside as acceptors. For transglycosylation reaction of stevioside, the transglycosylation rate was similar and the transglycosylation yield was increased compare with conventional process using liquefied starch as the donor. Also the accumulation of maltooligosaccharides in reaction mixture was minimized.

• 한국미생물·생명공학회지
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• 제26권2호
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• pp.187-194
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• 1998

수계에서 전분 가수분해효소의 transglycosylation반응을 이용하여 배당체(glycoside)를 합성하였다. Glycosyl donor인 starch와 glycosyl acceptor인 benzylalcohol을 반응기질로 선택하였다. 시판되는 9종의 당가수분해효소의 transglycosylation활성을 조사한 결과 glucose와 한 종류의 glycoside만을 생산하는 amyloglucosidase(from Rhizopus sp.)를 반응효소로 선정하였다. Amyloglucosidase에 의해 합성된 배당체는 여러 가지 분석을 통해 glucose의 1번 OH기에 benzylalcohol이 ${alpha}$형태로 결합된 benzylalcohol-${alpha}$-glucoside(BG)임을 확인하였다. 수계에서 이 효소에 의한 transglycosylation 반응의 최적조건은 starch 50mg/$m\ell$, benzylalcohol 50 mg/ml, 온도 45$^{\circ}C$, 효소량 10 unit/ml, pH 5.0, 반응시간 32시간이었으며 합성된 BG는 amyloglucosidase에 의해서는 분해되지 않았고 ${alpha}$-glucosidase에 의해 glucose와 benzylalcohol로 가수분해되었다.

• 한국식품영양과학회지
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• 제29권5호
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• pp.802-806
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• 2000

Cellobiose에 당전이 효소인 dextransucrase를 여러 가지 조건별로 반응시켜 올리고당의 생성경향을 알아보았다. Cellobiose에 대한 acceptor 반응의 최적조건은 cellobiose와 surcose의 비율은 3:1, 효소의 양은 2 U/mL, buffer의 이온강도는 25 mM, pH는 5, 반응온도는 $25^{\circ}C$로 나타났다. Cellobiose의 acceptor products는 종합도 6까지 생성되었으며, 구조는 2-O-isomaltodextrinyl cellobiose로 추정하였다.

• KSBB Journal
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• 제9권2호
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• pp.108-114
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• 1994

분쇄마찰매치 함유 불균일상 효소반응계에서 생전분을 당공여체로 한 stevioside의 당전이 반응의 kinetics에 관한 연구를 수행하였다. 생전분으로부터 CD를 생서하는 과정과 생성된 CD와 stevioside가 random sequenrial bireactant ,echanism으로 반응하여 당전이 equation을 유도하였다. 또한 유도된 반응식의 각종 kinetic constants을 평가하였다. 유도된 반응식을 Runge-Kutta integration법으로 계산하였으며, 계산 결과를 실험치와 비교하여 유도식의 효용성을 검토하였다. 유도된 kinetic equations는 당공여체인 생전분의 농도, stevioside의 농도, 그리고 중간산물인 CD의 농도 변화를 비교적 정획히 표시할 수 있었으며, 고효율 당전이 효소반응기 개발에 활용될 것이다.

• 미생물학회지
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• 제34권3호
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• pp.137-143
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• 1998

수계에서 전분 가수분해 효소의 transglycosylation 반응을 이용하여 배당체(glycoside)를 합성하였다. Glycosyl donor인 가용성전분과 glycosyl acceptor인 benzylalcohol을 기질로 하여 ${\alpha}$-amylase에 의해 합성되는 배당체는 glucose의 1번 OH기에 benzylalcohol이 ${\alpha}$형태로 결합한 benzylalcohol-${\alpha}$-glucoside(BG)와 benzylalcohol-${\alpha}$-maltoside(BM)이었다. pH 5.0의 반응에서는 주로 BG가, pH 8.0의 반응에서는 BM만이 합성되는 특이한 반응양상을 보였다. Transglycosylation 반응의 최적조건은 가용성전분 50 mg/ml, benzylalcohol 50 mg/ml, 온도 $30-35^{\circ}C$, 효소량 10 unit/ml이었으며, 합성된 BG는 ${\alpha}$-glucodisase에 의해 glucose와 benzylalcohol로 가수분해되었으며 BM의 경우는 pH 5.0에서는 glucose와 BG로 가수분해되었으나 pH 8.0이상에서는 전혀 분해되지 않았다. BM과 구조적으로 유사한 maltotriose는 pH 5.0에서 glucose와 maltose로 가수분해 되었으나 transglycosylation반응은 거의 일어나지 않았으며 pH 8.0에서는 가수분해도 transglycosylation반응도 일어나지 않았다.

• Journal of Microbiology and Biotechnology
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• 제18권5호
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• pp.901-907
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• 2008

Thermotoga neapolitana $\beta$-glucosidase (BglA) was subjected to site-directed mutagenesis in an effort to increase its ability to synthesize arbutin derivatives by transglycosylation. The transglycosylation reaction of the wild-type enzyme displays major ${\beta}(1,6)$ and minor ${\beta}(1,3)$ or ${\beta}(1,4)$ regioselectivity. The three mutants, N291T, F412S, and N291T/F412S, increased the ratio of transglycosylation/hydrolysis compared with the wild-type enzyme when pNPG and arbutin were used as a substrate and an acceptor, respectively. N291T and N219T/F412S had transglycosylation/hydrolysis ratios about 3- and 8-fold higher, respectively, than that of the wild-type enzyme. This is due to the decreased hydrolytic activity of the mutant rather than increased transglycosylation activity. Interestingly, N291T showed altered regioselectivity, as well as increased transglycosylation products. TLC analysis of the transglycosylation products indicated that N291T retained its ${\beta}(1,3)$ regioselectivity, but lost its ${\beta}(1,4)$ and ${\beta}(1,6)$ regioselectivity. The altered regioselectivity of N291T using two other acceptors, esculin and salicin, was also confirmed by TLC. The major transglycosylation products of the wild type and N291T mutant were clearly different. This result suggests that Asn-291 is highly involved in the catalytic mechanism by controlling the transglycosylation reaction.

• Bulletin of the Korean Chemical Society
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• 제19권11호
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• pp.1233-1238
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• 1998

Transglycosylation reactions among methyl 2,3,4,6-tetra-O-methyl-D-glycopyranosides and isomeric butyl alcohols or cyclohexanol took place in the presence of trimethylsilyl trifluoromethanesulfonate (TMSOTf) in dichloromethane. The extent of the reaction after 1 h and 24 h from mixing was determined by gas chromatography (GC). Anomerization of the substrate took place during the course of transglycosylation, which favors α anomer regardless of the anomeric configurations of the starting glycosides. Transglycosylation also favors the a anomer regardless of the steric bulkiness of the alcohol. tert-Butyl alcohol did not give any transglycosylation, suggesting the steric hindrance of approaching the bulky alcohol to the oxonium intermediate. A mechanism for the transglycosylation have been proposed.

• Journal of Microbiology and Biotechnology
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• 제16권11호
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• pp.1678-1683
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• 2006

A novel suspension-phase enzyme reaction system for the intermolecular transglycosylation of L-ascorbic acid into 2-O-${\alpha}$-D-glucopyranosyl L-ascorbic acid supplementing extrusion starch as the glycosyl donor was developed using cyclodextrin glucanotransferase from Thermoanaerobacter sp. A high conversion yield compared to the conventional soluble-phase enzyme reaction system using cyclodextrins and soluble starch was achieved. The optimal reaction conditions were 2,000 units of cycIodextrin glucanotransferase, 20 g/l of L-ascorbic acid, and 50 g/l of extrusion starch at $50^{\circ}C$ for 24 h. The new suspension-phase enzyme reaction system also exhibited several distinct advantages other than a high conversion yield, including a lower accumulation of oligosaccharides and easily separable residual extrusion starch by centrifugation or filtration in the reaction mixture, which will facilitate the purification of 2-O-${\alpha}$-D-glucopyranosyl L-ascorbic acid. The new suspension-phase enzyme reaction system seems to be potentially applicable as the industrial process for the production of thermally and oxidatively stable 2-O-${\alpha}$-D-glucopyranosyl L-ascorbic acid.

• 미생물학회지
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• 제36권1호
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• pp.8-13
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• 2000

Trichoderma koningii ATCC 26113에서 분비되는 효소인 $\beta$-glucosidase를 cellobiose, sophorose, laminaribiose 및 gentiobiose 등의 기질과 반응시킨 후 효소의 transglycosylation 반응 산물을 분석하였다. 각각의 기질로부터 생성된 이당체(dimer)들을 HPLC로 분리하고 $^(1)H$-NMR spectroscopy를 통하여 분석하였다. Cellobiose를 기질로 사용하여 효소와 반응시켰을 때 그 산물에는 laminaribiose, sophorose 및 gentiobiose가 포함되었음을 확인할 수 있었다. Laminaribiose, sophorose 및 gentiobiose를 기질로 사용하였을 경우에 효소는 transglycosylation 반응을 통하여 새로운 $\beta$-glycosidic 결합을 갖는 이당체들을 생성하였다. 효소반응에 의하여 누적되는 이당체의 양은 생성속도보다는 분해속도에 의하여 결정되는 것으로 나타났다.