Pullulanase를 생산하는 Aeromonas caviae No. S-76의 특성과 배양조건

손천배,김명희,이명자 충남대학교부설 생명공학연구소 1992 생물공학연구지 Vol.2 No.-

A bacterial strain No. S-76 which produced pullulanase powerfully was isolated from soil. The isolated bacterium was 0.4∼0.6×0.8∼1.4 ㎛ in size, gram negative, rods, motile and was identified as Aeromonas caviae by Bergey's manual of determinative bacteriology with various characteristics investigated. The highest yield of pullulanase of the strain was obtained by using the following medium: 1% pullulan, soluble starch or corn starch as a carbon sources and 0.5% yeast extract, peptone as nitrogen sources with an initial pH of 9.0. The optimal culture conditions for production of pullulanase were at 32℃ for 2 days.

Aeromonas caviae No. S-76이 생산하는 Pullulanase의 정제, 특성 및 Maltosyl-β-Cyclodixtrin의 합성

손천배,김명희,이명자 충남대학교부설 생명공학연구소 1992 생물공학연구지 Vol.2 No.-

The crude enzyme solution obtained by shaking culture of Aeromonas caviae No. S-76 isolated from soil as pullulanase producing bacterium was purified by 50 folds with 21% yield by salting out with ammonium sulfate and column chromatography using DEAE-Sephadex A-50 and Sephadex G-150. The purified pullulanase had a molecular weight of 118,000 approximately by SDS-polyacrylamide slab gel electrophoresis and pI of 4.3 by isoelectric focusing. And optimum reaction temperature and pH for pullulanase were 50℃ and 8.0, respectively. The purified enzyme was relatively stable at pH 6.0∼9.0 and below 45℃. This enzyme synthesized maltosyl-β-cyclodextrin from mixture of β-cyclodextrin and maltose.

Aspergillus niger 및 그 變異株의 生澱粉糖化酵素에 關한 硏究

孫天培,朴允仲 충남대학교 농업과학연구소 1983 農業技術硏究報告 Vol.10 No.1

Aspergillus niger IFO 8541 (NRRL 3112) was investigated through a series of UV rays and N-Methyl-N'-Nitro-N'-Nitrosoguanidine (NTG) treatments to induce mutants that produce highly active raw starch saccharifying enzyme, and two mutants with strong enzymatic productivity were obtained. The mutants obtained were investigated for their fungal characters, condition of enzyme production, and other activities. Furthermore, the raw starch saccharifying enzyme was purified and the characteristics of purified enzyme were studied. The results obtained were summarized as follows; 1. The color of conidial head of UV-46 mutant obtained from UV rays treatment was changed to tan type and the gelatinated starch saccharifying enzyme productivity and the raw starch saccharifying enzyme productivity increased up to twice and 1.8 times compared to the productivities of original Aspergillus niger IFO 8541 cultured on the wheat bran, respectively. 2. The conidial head color of NG-41 mutant obtained from NTG treatment became lighter than that of parent strain. The gelatinated starch saccharifying enzyme productivity and raw starch saccharifying enzyme productivity increased about 1.8 times, and twice over the Aspergillus niger I Fo 8541 parent strain cultured on wheat bran, respectively. The productivity of α-amylase increased about 3 times more than the parent strain. 3. Two peaks of glucoamylase and a peak of α-amylase were obtained when enzyme solution of mutants and parent strain were passed through DEAE-Sephadex A-50 column chromatography. Glucoamylase I showed only gelatinated starch saccharifying enzyme activity. However, glucoamylase Ⅱ(raw starch saccharifying enzyme) showed both raw starch saccharifying enzyme activity and gelatinated starch saccharifying enzyme activity. 4. Mutant, UV-46 was strengthened in glucoamylase Ⅱ productivity and mutant, NG-41 was strengthened in α-amylase productivity. 5. Glucoamylase Ⅱ of mutants and parent strain were appeared to have the same enzymatic properties. 6. Glucoamylase Ⅱ of mutants and parent strain were recognized as simple enzyme through electrophoresis. 7. The glucoamylase Ⅱ crystallized showed rhombic board type. 8. The molecular weight, isoelectric point, optimum pH, and optimum temperature of the glucoamylase Ⅱ crystallized were estimated as 76,000, 3.4, 3.5 and 60℃, respectively.

黑麴菌의 變異에 있어서 生澱粉糖化 酵素活性의 變動

孫天培 충남대학교 자연과학연구소 1987 忠南科學硏究誌 Vol.14 No.1

This study was to induce mutant irradiated with UV rays from Soju brewing black-Koji molds-Aspergillus awamori that produces highlv raw starch saccharyfying enzymes, and the obtained mutant was investigated for fungal characters, condition of enzyme production, characteristics of purified enzyme. The mutant strain showed morphological characteristics such as smaller conidiospore, vesicle, conidial head, and lighter conidia compared with the parent strain. Also when it was cultivated in wheat bran media, the enzyme productivity was beat 2-3 days at 30℃. The raw starch saccharifying enzyme activity was increased approxymately two times and acid protease activity was increased seven times. By the enzyme purification, two glucoamylases were separated. The peak Ⅰ showed gelatinized starch saccharifying activity but the peak Ⅱ showed both raw and gelatinized starch saccharifing activity. The molecular weight, and the optimum pH of the raw starch saccarifying enzyme were 94,000, 3.5, and it was stable at pH 2-9. The amount of sugars in the carbohydrate - protein link - age was 17% and they were identified as mannose, glucose, and galactosamine. When various kinds of raw starches were saccharified by using enzyme which had same gelatinized starch saccharifing activity, it was found that the mutant strain saccharified faster than the parent strain. The saccharifying speed of the parent enzyme was in the order of corn>wheat>cassava>sweet potato>potato starch, but in the mutant strain enzyme, cassava starch saccharified faster than wheat starch. The acid protease produced from the mutant strain did not modify with raw starch saccharifying enzyme and it did not affect enzyme activity.

호알칼리성 Bacillus sp. No. 4의 Cyclodextrin Glycosyltransferase에 의한 Glycosyl Sucrose의 생산과 저충치성 당으로서의 응용

손천배,유미경,김명희,문숙경 충남대학교부설 생명공학연구소 1992 생물공학연구지 Vol.2 No.-

Action of a cyclodextrin glycosyltransferase (CGTase) produce from alkalophilic Bacillus sp. No. 4 was studied in a solution containing starch and sucrose to prepare glycosyl sucrose syrup with good sweetness and antidecaying properties of teeth. In the initial stage of the reaction the CGTase produced cyclodextrin, however, the cyclodextrin disappeared and glycosyl sucrose was formed with the lapse of reaction time. The best proportion of sucrose to starch for prodution of glycosyl sucrose was about 1 : 1. The optimum pH and temperature of the coupling reaction was pH 6.0 and 60℃, respectively. Main composition of glycosyl sucrose syrup prepared with 20% starch and 20% sucrose was sucrose 18%, glucosyl sucrose (G_2F) 15.3% and amltosyl sucorse (G_3F) 11.3%. And glucose, maltose and maltotriose were produced very little. Smaller amounts of acid and insoluble glucan were formed in the syrup by Streptococcus mtans OMZ176 than in the sucrose. Therefore, the prepared glycosyl sucrose sucrose syrup is expected to prevent teeth from decaying.

Candida muscorum의 Protease生産條件과 그 性質에 關한 硏究

孫天培 충남대학교 대학원 1974 論文集 Vol.3 No.-

A yeast was selected as a potent producer of an extracellular protease and identified by J.Lodder's manual and it was investigated cultural conditions for protease production in wheat bran medium and properties of its protease. The results obtained were as follows. 1. The sellected strain was identified as a strain belonging to Candida muscorum. 2. The optimum conditions for protease production in wheat bran media were: water content added 75%, temperature 25℃, incubation time 4days, initial pH 6.0 and the thickness of medium 1.2㎝. 3. No significant effect was found in the case of the addition of various bran and cake to wheat bran medium. 4. When 2.5% of sucrose added to wheat bran medium, the protease production was increased about 40%, when 0.5% of (NH_(4))_(2)SO_(4) added to wheat bran medium, the protease production was increased about 30%. 5. The protease of selected strain was divided into the neutral protease having maxilmum activity at pH6.0 and acid protease having maximum activity at pH 3.5. 6. The protease of selected strain showed maximum activity at 50℃, when reaction was proceeded in 3% milk casein solution for 3hrs. 7. The protease activity in crude enzyme solution was decreased 25% during treated for 15 minutes at 50℃ and was decreased 88% during treated for 15 minutes at 60℃.

Aeromonas caviae No. S-76이 생산하는 Pullulanase의 정제, 특성 및 Maltosyl-β-Cyclodextrin의 합성

손천배,김명희,이명자 한국산업미생물학회 1991 한국미생물·생명공학회지 Vol.19 No.4

Pullulanase 생산균으로서 토양으로부터 분리한 Aeromonas caviae No. S-76을 진탕배양하여 얻은 조효소액을 ammonium sulfate 침전, DEAE Sephadex A-50 column chromatography, Sephadex G-150 column chromatography에 의하여 정제하였다. 이때 수율은 21%이었고 50배의 정제도를 가진 효소단백질을 얻었다. 정제효소는 SDS-polyacrylamide slab gel 정기영동에 의하여 분자량 118,000의 단일단백질이었고, 등전점은 4.3, 작용 최적온도는 50℃, 작용 최적 pH는 8.0이었다. 또한 이 효소는 45℃ 이하, pH 6.0∼9.0 범위에서 안정성을 나타내었다. 이 효소를 β-cyclodextrin과 maltose의 고농도 혼합액에 작용시켜 maltosyl-β-cyclodextrin을 합성하였다. The crude enzyme solution obtained by shaking culture of Aeromonas caviae No. S-76 isolated from soil as pullulanase producing bacterium was purified by 50 folds with 21% yield by salting out with ammonium sulfate and column chromatography using DEAE-Sephadex A-50 and Sephadex G-150. The purified pullulanase had a molecular weight of 118,000 approximately by SDS-polyacrylamide slab gel electrophoresis and pI of 4.3 by isoelectric focusing. And optimum reaction temperature and pH for pullulanase were 50℃ and 8.0, respectively. The purified enzyme was relatively stable at pH 6.0∼9.0 and below 45℃. This enzyme synthesized maltosyl-β-cyclodextrin from mixture of β-cyclodextrin and maltose.

Pullulanase를 생산하는 Aeromonas caviae No. S-76의 특성과 배양조건

손천배,김명희,이명자 한국산업미생물학회 1991 한국미생물·생명공학회지 Vol.19 No.4

Pullulanase 생산력이 높은 세균 No. S-76을 토양으로부터 분리하였다. 분리된 균은 0.4∼0.6×0.8∼1.4㎛의 크기의 gram음성, 간균으로서 운동성이 있으며, 여러 가지 특성을 조사한 결과 Bergey의 세균분류 동정법에 따라 Aeromonas caviae로 동정되었다. 본 균의 pullulanase 생산배지로서는 탄소원은 1% pullulan, soluble strach 또는 corn starch가, 질소원으로는 0.5% yeast extract 또는 peptone이 적당하였으며 initial pH는 9.0의 배지가 가장 좋았으며 32℃에서 2일간 배양이 적당하였다. A bacterial strain No. S-76 which produced pullulanase powerfully was isolated from soil. The isolated bacterium was 0.4∼0.6×0.8∼1.4㎛ in size, gram negative, rods, motile and was identified as Aeromonas caviae by Bergey's manual of determinative bacteriology with various characteristics investigated. The highest yield of pullulanase of the strain was obtained by using the following medium: 1% pullulan, soluble starch or corn starch as a carbon sources and 0.5% yeast extract, peptone as nitrogen sources with an initial pH of 9.0. The optimal culture conditions for production of pullulanase were at 32℃ for 2 days.

Cyclodextrin Glycosyltransferase 를 생산하는 호 Alkali 성 Bacillus sp. No. 4. 의 특성과 배양조건

손천배(Sheon Bae Sohn),문숙경(Suk Keung Moon),이근희(Keun Hoi Lee) 충남대학교 생활과학연구소 1990 忠南生活科學硏究誌 Vol.3 No.-

호알칼리성 Bacillus sp. No. 4의 Cyclodextrin Glycosyltransferase에 의한 Glycosyl Sucrose의 생산과 저충치성 당으로서의 응용

손천배(Cheon-Bae Sohn),유미경(Mi-Kyeong You),김명희(Myung-Hee Kim),문숙경(Suk-Keung Moon) 한국식품과학회 1991 한국식품과학회지 Vol.23 No.4