• 한국동물학회:학술대회논문집
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• 한국동물학회 2001년도 한국생물과학협회 학술발표대회
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• pp.152.1-152
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• 2001

• Journal of Applied Biological Chemistry
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• 제51권6호
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• pp.292-295
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• 2008

Three kinds of oligosaccharides were obtained from the hydrolysate of brown copra meal galactomannan by a purified extracellular ${\beta}$-mannanase from Trichoderma sp. These oligosaccharides were identified as Man-Man, ${Gal^2}{Man_3}(6^2 mono-O-{\alpha}-D-galactopyranosyl-4-O-{\beta}-D-mannotriose)$, and ${Gal^2}{Man_6}(6^2-mono-O-{\alpha}-D-galactopyranosyl-4-O-{\beta}-D-mannohexaose)$, where Gal- and Man-represent ${\alpha}$-1,6-D-galactosidic and ${\beta}$-1,4-mannosidic linkages, respectively. The mode of action of ${\beta}$-mannanase on brown copra meal galactomannan is described on the basis of the structure of these oligosaccharides.

• 한국미생물·생명공학회지
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• 제26권4호
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• pp.316-322
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• 1998

N-$\varepsilon$-aminocaproyl-$\alpha$-D-galactopyranosylamine-sepharose를 담체로 하는 affinity chromatography에 의한 해바라기씨 유래 $\alpha$-galactosidase($\alpha$-D-galactoside galactohydrolase EC 3. 2. 1. 22)의 정제방법과 정제효소에 대한 효소화학적 성질을 규명하였다. N-$\varepsilon$-aminocaproyl-$\alpha$-D-galactopyranosylamine의 흡착제를 합성하여 sepharose에 coupling하였다. 기질 p-nitrophenyl $\alpha$-D-galactopyranoside에 대한 정제효소의 비활성은 291.66 units/mg였고, 조효소와 비교하여 115배의 정제 배율을 나타내었다. 정제효소의 순도는 SDS-polyacryl amide gel전기 영동법 에 의해 단일 band를 나타내었으며, 분자량은 42,000으로 추정되었다. 정제효소의 최적 pH와 온도는 4.5, 55$^{\circ}C$이며, pH 4-5, 30-55$^{\circ}C$의 범위에서 pH와 온도 안정성을 나타내었다. 또한 정제효소는 Ag$^{2+}$, Hg$^{2+}$, CO$^{2+}$의 금속에 의해 70%이상의 저해효과를 나타내었다. 정제효소는 melibiose, raffinose 및 copra galactomannan에 대한 galactose의 유리를 TLC에 의해 확인하였고, 각 기질에 대한 galactose의 가수분해 속도를 HPLC에 의해 비교하였다.

• 한국식품과학회지
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• 제52권6호
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• pp.595-603
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• 2020

토란에 존재하는 점질다당의 새로운 이용방안을 모색하기 위하여 토란으로부터 다당을 분리하여 항보체 활성을 평가하고 구조 분석을 행하였다. 토란으로부터 분리한 조다당 CE를 이온교환수지와 Sephadex G-100 column를 이용하여 정제하였고, 그 중 수율과 활성이 양호한 CE-4a를 최종 획분으로 선정하였다. 초기 면역반응에 중요한 역할을 하는 보체계에 대한 토란 다당의 활성화 여부를 측정한 결과 양성대조군인 PSK에 준하는 강력한 항보체 활성을 보였고, 시료의 농도차이를 두어 실험한 결과 농도 의존적임을 알 수 있었다. CE-4a는 분자량 약 182.4 kDa의 다당체로 구성당 조성을 확인한 결과 Man, Gal 및 GalA를 높은 비율로 함유하고 있었다. 본 당쇄의 결합양식을 규명하기 위하여 methlyation analysis를 행한 결과 CE-4a는 terminal Galp. 3-linked-Galp, 4-linked Manp, 2,4,6-linked Manp를 포함한 총 10종의 결합으로 구성되어 있었다. 또한 CE-4a의 전체구조를 추정하기 위하여 endo-α-(1→4)-polygalacturonase, exo-α-galactosidase 및 endo-β-(1→4)-mannanase를 이용한 연속 가수분해 처리 및 해석을 행하였다. 결과를 종합하면, 토란 유래 다당 CE-4a는 (1→4)-mannan 주쇄로 존재하며 주쇄인 mannose의 C(O)6 위치에서 한가닥 또는 C(O)2, C(O)6 위치에서 동시에 두가닥의 측쇄가 연결되어 존재하고, 측쇄는 주로 galacto oligo당이 측쇄로 분지된 특징이 있음을 확인할 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제6권2호
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• pp.86-91
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• 1996

Thermostable $\beta$-mannanase from Bacillus sp. YA-14 was purified by acetone precipitation, CM-cellulose, Sephadex G-100 and hydroxyapatite column chromatography from culture supernatant. The final enzyme preparation appeared to be homogeneous on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). $\beta$-Mannanase appeared to be a monomeric protein with a molecular weight of 67, 000 daltons. The optimal pH and temperature of the enzyme reaction were pH 6.0 and $75^{\circ}C$ , respectively. The enzyme was stable at a pH range of 6.0 to 9.0 and at temperatures between 45 and $85^{\circ}C$. The kinetic constants of $\beta$-mannanase as determined with a galactomannan (locust bean) as substrate were a Vmax of 25 unit/ml and a Km of 1.1 mg/ml. The enzyme had only limited activity on galactomannan substrate. It was suggested that mg $\beta$-mannanase activity is limited by the number of branched $\alpha$-galactose residues.

• 한국미생물·생명공학회지
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• 제27권4호
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• pp.298-303
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• 1999

An $\alpha$-D-galactosidase ($\alpha$-D-galactoside galactohydrolase, EC 3. 2. 1. 22) from earthworm was purified by affinity chromatography using N-$\varepsilon$-aminocaproyl-$\alpha$-D-galactopyranosylamine coupled to sepharose and its properties were examined. The specific activity of the purified enzyme, tested with p-nitrophenyl-$\alpha$-D-galactopyranoside as substrate, was 314 units/mg protein, representing an 122-fold purification of the original crude extract. The final preparation obtained from by Sephadex G-25 chromatography showed a single band on SDS-polyacrylamide gel electrophoresis. The molecular weight was determined to be 48,000 by SDS-polyacrylamide gel electrophoresis. The purified galactosidase was showed maximum activity at pH 4.5 and 4$0^{\circ}C$, and was stable in the pH and temperature ranges from 4.0 to 5.5 and 30 to 5$0^{\circ}C$, respectively. The enzyme activity was inhibited by Zn2+, Hg2+ and Co2+. When the purified $\alpha$-galactosidase treated to guar gum for 6 hour, gel-promoting property was increased. It was clear that enzymatic elimination of galactose from guar gum by purified $\alpha$-galactosidase would lead to a significant increase in gelation ability.

• Journal of Microbiology and Biotechnology
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• 제1권2호
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• pp.90-95
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• 1991

A ${\alpha}-galactosidase{\;}({\alpha}-D-galactoside$ galactohydrolase; EC 3.2.1.22) was purified from the culture filtrate of Penicillium purpurogenum by DEAE-cellulose column chromatography, gel filtration of Bio gel p-l00, and subsequent SP-Sephadex C-25 chromatography. The final preparation thus obtained showed a single band on polyacrylamide disc-gel and SDS-polyacrylamide gel electrophoresis. The molecular weight and isoelectric point were determined to be 63,000 and pH 4.0 by SDS-polyacrylamide gel electrophoresis and isoelectric focusing, respectively. The galactosidase exhibited maximum activity at pH 4.5 and $55^{\circ}C$, and was stable between pH 2 and 5, and also stable up to $40^{\circ}C$. The enzyme activity was not affected considerably by treatment with other metal compounds except mercuric chloride and silver nitrate. Copra galactomannan was finally hydrolyzed to galactose, mannose and mannobiose through the sequential actions of the purified galactosidase and mannanase from the same strain. The enzyme hydrolyzed melibiose and raffinose, but not lactose.

• Journal of Microbiology and Biotechnology
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• 제24권8호
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• pp.1044-1050
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• 2014

Since there is no consensus about the most reliable assays to detect invasive aspergillosis from samples obtained by minimally invasive or noninvasive methods, we compared the efficacy of an enzyme-linked immunosorbent assay (ELISA) for galactomannan (GM) detection and quantitative real-time PCR assay (qRT-PCR) for the diagnosis of invasive pulmonary aspergillosis. Neutropenic, male Sprague-Dawley rats (specific pathogen free; 8 weeks old; weight, $200{\pm}20g$) were immunosuppressed with cyclophosphamide and infected with Aspergillus fumigatus intratracheally. Tissue and whole blood samples were harvested on days 1, 3, 5, and 7 post-infection and examined with GM ELISA and qRT-PCR. The A. fumigatus DNA detection sequence was detected in the following number of samples from 12 immunosuppressed, infected rats examined on the scheduled days: day 1 (0/12), day 3 (0/12), day 5 (6/12), and day 7 (8/12) post-infection. The sensitivity and specificity of the qRT-PCR assay was 29.2% and 100%, respectively. Receiver operating characteristic curve (ROC) analysis indicated a Ct (cycle threshold) cut-off value of 15.35, and the area under the curve (AUC) was 0.627. The GM assay detected antigen in sera obtained on day 1 (5/12), day 3 (9/12), day 5 (12/12), and day 7 (12/12) post-infection, and thus had a sensitivity of 79.2% and a specificity of 100%. The ROC of the GM assay indicated that the optimal Ct cut-off value was 1.40 (AUC, 0.919). The GM assay was more sensitive than the qRT-PCR assay in diagnosing invasive pulmonary aspergillosis in rats.

• Journal of Applied Biological Chemistry
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• 제53권2호
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• pp.71-76
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• 2010

가정에서 제조된 된장으로부터 분리된 Bacillus licheniformis WL-12의 mannanase 유전자를 크로닝하여 그 염기서열을 결정한 결과 mannanase 유전자는 360 아미노산으로 구성된 단백질을 코드하며 1,080 뉴클레오티드로 이루어졌다. 아미노산 잔기배열을 분석한 결과 WL-12의 mannanase는 GH family 26에 속하는 B. licheniformis DSM13의 mannanase와 동일하였다. B. lichenifromis WL-12의 mannanase 유전자를 함유한 재조합대장균의 균체파쇄상등액으로부터 부분정제된 효소를 사용하여 반응특성을 조사하였다. pH 6.0과 $65^{\circ}C$에서 최대 반응활성을 보였으며, locust bean gum (LBG)과 konjac의 분해능은 높으나 guar gum의 분해능은 낮았다. Mannanase로 LBG와 mannooligosaccharides를 분해하였을 때 mannose, mannobiose와 mannotriose가 주된 최종 반응산물로 관찰되었으며 mannobiose는 분해하지 못하였으나 이보다 중합도가 큰 mannooligosaccharides은 분해하였다.

• 미생물학회지
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• 제46권4호
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• pp.397-400
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• 2010

만난의 분해활성을 갖는 Paenibacillus woosongensis의 배양 상등액으로부터 mannanase, ${\beta}$-mannosidase와 ${\alpha}$-galactosidase 활성이 관찰되었다. 배양상등액 내의 locust bean gum를 분해하는 mannanase는 pH 5.5와 $60^{\circ}C$, para-nitrophenyl-${\beta}$-D-mannopyranoside를 분해하는 ${\beta}$-mannosidase는 pH 6.5, $50^{\circ}C$, para-nitrophenyl-${\alpha}$-D-galactopyranoside를 분해하는 ${\alpha}$-galactosidase는 pH 6.0-6.5와 $50^{\circ}C$에서 각각 최대활성을 보였다. 배양상등액의 만난 분해효소는 mannotriose, mannotetraose, mannopentaose, mannohexaose와 같은 만노올리고당을 분해할 뿐 아니라 mannobiose도 분해하였다. 또한 melibiose, raffinose, stachyose 등의 ${\alpha}$-1,6 결합형 galacto-oligosaccharides를 분해하여 galactose를 생성하였다. 이러한 결과로 보아 P.woosongensis는 galactomannan을 완전히 분해하는데 필요한 3종류 효소를 모두 생산하는 것으로 판단된다.