• 한국미생물·생명공학회지
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• 제20권2호
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• pp.156-163
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• 1992

Cyclodextrin glucanotransferase 생산균주 선발배지를 이용하여 국내 토양으로부터 CGTase 활성이 우수한 Bacillus sp. E1균주를 분리하였다. FPLC를 이용하여 gel filtration과 anion exchange column chromatography를 한 결과 순수 정제된 단일 단백질을 얻을 수 있었으며, 정제된 효소의 최적 작용 pH 범위는 6에서 8까지 였고, 온도는 $60^{\circ}C$에서 최적을 나타냈다. 정제된 단백질의 분자량은 114,000, 등전점은 4.3이었다. 생성된 cyclodextrin은 $\beta$와 $\gamma$-cyclodextrin이 주였으며, 특이하게도 $\alpha$-cyclodextrin은 거의 생성되지 않았다. 작용 후 25시간 후 최대의 $\beta$-cyclodextrin이 생성되었으며, 이때 $\beta$-cyclodextrin과 $\gamma$-cyclodextrin의 생성비율은 7:1이었다.

• 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.

• Journal of Microbiology and Biotechnology
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• 제11권2호
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• pp.242-250
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• 2001

A Gram-positive bacterium (strain JB-13) that was isolated from soil as a producer of cyclodextrin glucanotransferase (CGTase) [EC 2.4.1.19] was identified as Panibacillus sp. JB-13. This CGTase could catalyze the transglucosylation reaction from soluble starch to L-ascorbic acid (AA). A main product formed by this enzyme with ${\alpha}-glucosidase$ was identified as $2-O-{\alpha}-D-glucopyranosyl{\;}_{L}-ascorbic$ acid (AA-2G) by the HPLC profile and the elemental analysis. CGTase was purified to homogeneity using ammonium sulfate fractionation, ion-exchange chromatography on DEAE-Seohadex A-50, and gel chromatography on Sephacryl S-200HR. The molecular weight was determined to be 66,000 by both gel chromatography and SDS-PAGE. The isoelectric point of the purified enzyme was 5.3. The optimum pH and temperature was PH 7.0 and $45^{\circ}C$ respectively. The enzyme was stable in the range of pH 6-9 and at temperatures of $75{\circ}C$ or less in the presence of 15 mM ${CaCl_2}.\;{Hg^2+},\;{Mn^+2},{Ag^+},\;and\;{Cu^2+}$ all strongly inhibited the enzyme's activity.

• Bulletin of the Korean Chemical Society
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• 제24권11호
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• pp.1627-1631
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• 2003

Molecular modeling was performed to comprehend the chiral recognition of ${\alpha}$-methoxy-${\alpha}$-trifluoromethylphenylacetic acid (MTPA) enantiomers by cyclomaltoheptaose (${\beta}$-cyclodextrin,${\beta}$-CD) and 6-amino-6-deoxy-cyclomaltoheptaose (am-${\beta}$-CD). Monte Carlo (MC) docking coupled to constant temperature molecular dynamics (MD) simulations was applied to the investigation for the ${\alpha}$-methoxy-${\alpha}$-trifluoromethylphenylacetic acid complexation with two different CDs in terms of the relative distribution of the interaction energies. The calculated results are finely correlated with the experimental observations in chiral recognition thermodynamics. Am-${\beta}$-CD as a host showed the superior enantio-discrimination ability to the native ${\beta}$-CD where the amino group of am-${\beta}$-CD was critically involved in enhancing the ability of chiral discrimination via the Coulombic interaction with MTPA.

• Biotechnology and Bioprocess Engineering:BBE
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• 제3권2호
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• pp.78-81
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• 1998

Cyclodextrin glucanotransferase [CGTase, E.C.2.4.1.19] is an extracellular enzyme, which catalyzes he formation of ${\alpha}$-, ${\beta}$-, ${\gamma}$- CDs from starch. Their proportions of formations depend on enzyme sources and reaction conditions. To understand what determines the product specificity of CGTases, we examined the alteration of product specificity of CGTase from Bacillus macerans by organic solvent sand pH. At acidic pH range less than pH 6 where the enzyme was unstable, the ratio of ${\alpha}$-/ ${\beta}$-CD production was increased 4 times more than that at neutral pH range. As we increased the concentration of 2-butanol, ${\alpha}$-/ ${\beta}$-CD ratio was proportionally increased but / ratio remained constant. The ${\alpha}$-/ ${\beta}$-CD ratio of products was increased in the reaction media which yielded low products.

• 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.

• KSBB Journal
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• 제10권5호
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• pp.491-498
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• 1995

Cyclodextrin의 고순도 분획, 정제를 위한 CD adsorbent의 제조에 척합한 matnx를 선별하기 위하여 capric acid를 ligand로 각종 이온교환수지를 비교 검토한 결과 DEAE Cellulose가 가장 적합함을 알았다. DEAE Cellulose와 capric acid간의 결합 안정성은 온도, ionic strength의 변화에 영향을 받았으며, ethanol 농도의 변화에는 안정하였다. CD adsorbent의 흡착량, 탈착량, 그리고 ${\alpha}$-, ${\beta}$- 및 ${\gamma}$-CD 의 용출양상을 규명하였다. 탄소쇄가 다른 각종 포화, 불포화 fatty acid를 ligand로 하여 specific adsorbent를 제조하였으며, ${\alpha}$-, ${\beta}$- 및 ${\gamma}$-CD의 회수율과 분리능을 비교 검토 하였다. 그 결과 steanc a acid는 ${\alpha}$-CD 그리고 linoleic acid는 ${\beta}$-CD에 대하여 높은 회수율과 선택성을 보였으며, 선택성은 fatty acid의 탄소쇄 길이와 이중결합 유무에 의하여 영향을 받았다. Stearic acid와 linoleic acid CD adsorbents를 이용하여 CD 혼합물로부터${\alpha}$-, ${\beta}$- 및 ${\gamma}$-CD의 분획양상을 검토하였다.

• 한국식품과학회지
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• 제16권4호
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• pp.398-402
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• 1984

Rhizopus oryzae 가 생산(生産)하는 glucoamylase 의 각종기질(各種基質)에 대(對)한 분해반응(分解反應)을 검토(檢討)하였다. Glucoamylase I 과 II 는 amylose, amylopectin, glycogen, 가용성 전분, pullulan, maltose, maltotriose, maltotetriose, maltopentaose, maltohexaose, maltoheptaose, maltooctaose를 가수분해(加水分解)하였으나, ${\alpha}-cyclodextrin$, ${\beta}-cyclodextrin$, sucrose, raffinose, 젖당은 가수분해(加水分解)하지 못하였다. $37^{\circ}C$, 32시간(時間)의 반응(反應)에서 glucoamylase I 은 amylopectin, 가용성 전분, amylose를 거의 100% 분해하였고 glycogen 만을 88%정도 분해 하였으나, glucoamylase II 는 공시기질(供試基質) 4 종(種)을 거의 100% 분해하였다. Glucoamylase I 과 II 의 반응생성물질(反應生成物質)은 glucose 만이었고 ${\alpha}-glucosyltransferase$ activity는 없었다. Glucoamylase I 과 II 는 생찹쌀 전분을 가장 잘 분해(分解)시키나 생감자 전분, 생미숙 바나나 전분, 생칡 전분, 생참마 전분, raw high amylose corn starch의 분해능(分解能)은 glucoamyase I 에 비(比)하여 생전분(生澱粉)의 분해력(分解力)이 강(强)하였다.

• Journal of Pharmaceutical Investigation
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• 제24권2호
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• pp.85-94
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• 1994

Inclusion complexes of ketoconazole (KT) with ${\alpha}-$, ${\beta}-cyclodextrin$ (CD) and dimethyl-${\beta}-cyclodextrin$ $(DM{\beta}CD)$ in a molar ratio of 1:2 were prepared by freeze-drying and solvent evaporation methods. The interactions of KT with ${\alpha}-CD$, ${\beta}-CD$ and $DM{\beta}CD$ in aqueous solution and in solid state were investigated by solubility study, infrared (lR) spectroscopy and differential scanning calorimetry (DSC). The stability constant of $KT-DM{\beta}CD$ inclusion complex (lC) was found to be the largest among three inclusion complexes. Clear differences in IR spectra and DSC curves were observed between inclusion complexes and physical mixtures (PM) of KT-CDs. It was also shown by IR spectra and DSC curves that solvent evaporation method might be. superior to the freeze-drying method in preparing the inclusion complexes of KT-CDs. The dissolution rate of KT was markedly increased by inclusion complex formation with CDs in the buffer solution at pH 4.0 and pH 6.8. The mean dissolution time (MDT,min), which represents the rapidity of dissolution, was in the order of $KT-DM{\beta}CD$ IC (3.20) < $KT-{\beta}-CD$ IC (4.36) < $KT-{\alpha}-CD$ IC (6.99) < $KT-{\alpha}-CD$ PM (17.46)< $KT-{\beta}-CD$ PM (19.36) < $KT-{\beta}-CD$ PM (28.53). The dissolution rates of KT-CD ICsprepared by solvent evaporation method were faster than those of KT-CD ICs prepared by freeze-drying method.

• Preventive Nutrition and Food Science
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• 제7권3호
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• pp.310-316
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• 2002

Extracellular cyclodextrin glucanotransferase (CGTase) from Paenibacillus sp. JK-12 was purified through sev-eral purification steps consisting of ammonium sulfate precipitation and chromatographies on DEAE-sephadex A-50 and Mono QIM HR5/5. The purified CGTase exhibited a single band on SDS-PAGE and was estimated to be approximately 82 kDa. The isoelectric point of the enzyme was 7.2 as determined by isoelectric focusing. The CGTase from Paenibacillus sp. JK-12 had a transglucosylation activity at the C-2 position of L-ascorbic acid. The optimum pH and temperature for the CGTase activity were 8.0 and 5$0^{\circ}C$, respectively. The enzyme activity was stable from pH 6.0 to 9.() and at temperatures up to 55$^{\circ}C$ at pB 8.0, having 80% residual activity. The activity of the CGTase was strongly resistant to metals such as A $g^{+}$ and $Ba^{2+}$ but slightly inhibited by H $g^{+}$, N $i^{2+}$ and $Mg^{2+}$. The enzymeproduced $\alpha$ -cyclodextrin ($\alpha$-CD) and $\beta$-CD as the main products from starch, but not ${\gamma}$-CD.X>-CD.