• 한국미생물·생명공학회지
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• 제19권6호
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• pp.592-597
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• 1991

토양 분리균인 B.stearothermophilus가 생산하는 xylanase를 ammonium sulfate 분획, DEAE-Sepharose CL-6B ion exchange chromatography, Sephadex G-100 gel 여과 및 열처리 등의 과정을 거쳐 단일 단배질로 분리 정제하였다. 170,000의 분자량을 본 정제 xylanase는 pH8과 pH10 사이의 넓은 최적 pH를 보였으며 $55^{\circ}C$에서 최대 활성을 나타내었다. $55^{\circ}C$에서 2시간의 열처리에 의해서도 활서의 손실이 거의 없을 정도로 열에 매우 안정하였으며 $co^{2+}$와 $Mn^{2+}$에 의해서 현저한 활성화 효과를 보였다.

• Asian-Australasian Journal of Animal Sciences
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• 제3권2호
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• pp.115-120
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• 1990

Extracellular nuclease(s) in buffalo rumen fluid were purified from strained rumen fluid by a procedure involving Seitz filtration, acetone fractionation and gel filtration on Sephadex G-100. The enzyme resolved into two peaks exhibiting both DNase and RNase activities. The molecular weight of enzyme corresponding to peaks I and II were approximately 30,000 and 12,000 respectively. The properties of enzymes from the two peaks, however, were same. Optimum temperature for both DNase and RNase activities was at $50^{\circ}C$. Whereas DNase activity was stable upto $60^{\circ}C$, RNase activity was stable only up to $50^{\circ}C$. DNase activity recorded two pH optima, one at pH 5.5 and the other at pH 7.0. RNase activity recorded a broad pH optimum between pH 6.0-8.0. pH stability of the enzyme coincided with pH optima for both the activities. DNase activity was stimulated by $Mg^{2+}$ and $Mn^{2+}$ and inhibited by $Fe^{2+}$, $Zn^{2+}$, $Hg^{2+}$ and $Ag^+$. RNase activity was also stimulated by $Mg^{2+}$ and $Mn^{2+}$ and inhibited by $Cu^{2+}$, $Fe^{2+}$, $Zn^{2+}$, $Hg^{2+}$ and $Ag^+$. Reducing agents stimulated both the activities.

• Journal of Microbiology and Biotechnology
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• 제13권5호
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• pp.794-799
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• 2003

Enzyme-linked immunosorbent assay (ELISA) was developed for the rapid detection of Fusarium species, known as harmful fungi in food. One of the hybridoma cell lines (lB8) which produced a monoclonal antibody (Mab) specific to Fusarium extracellular polysaccharide (EPS) was screened and the Mab was produced and purified. A detection limit of the sandwich ELISA against F. moniliforme EPS was $0.001\;\mu\textrm{g}/ml$ in the standard curve. Among the 59 strains tested, most Fusarium species showed hight reactivity with Mab lB8, even when the culture broths were diluted 100,000 times. On the other hand, the other genera, except A. versicolor and Trichoderma viride, had no reactivity. When 1 to $100\;\mu\textrm{g}$ of F. moniliforme EPS was spiked into rice, potato, and mandarine orange, the average recoveries were 151%, 84%, and 94%, respectively, determined by sandwich ELISA. The correlation coefficients between the EPS levels determined by sandwich ELISA and the dry mycelial weight of the liquid culture of F. moniliforme, as well as between the EPS and colony forming unit in solid culture of potato, were 0.97 and 0.91, respectively.

• Journal of Microbiology and Biotechnology
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• 제9권6호
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• pp.764-772
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• 1999

To develop an enzyme-linked immunosorbent assay (ELISA) for detecting mold, we produced anti-mold polyclonal antibodies by immunizing extracellular polysaccharide (EPS) of Aspergillus flavus or Penicillium citrinum in rabbits subcutaneously. Using the purified antibody (Ab) and Ab-HRP conjugate, a sandwich ELISA for EPS was established. The standard curve of the ELISA showed the detection limit for P citrinum EPS to be $0.003{\;}\mu\textrm{g}/ml$. The cross-reactivities of the anti-P citrinum EPS Ab toward components of P citrinum such as EPS, liquid, and solid culture mycelium were 100, 10.5, and 0.58%, respectively, and those toward components of A. flavus such as EPS, liquid and solid culture mycelium, and spore were 300, 0.67, 0.29, and 0%, respectively. When the reactivities toward culture broths of 59 mold strains were tested by the sandwich ELISA, most of the Aspergillus (16 of 18) and Penicillium (14 of 16) strains along with one of the two Cladosporium strains gave positive signals in the culture broths even when diluted 1,000 fold, while the rest of species such as Fusarium, Absidia, Alternaria, and Candida gave negative signals. When the water extracts of 30 corn samples were analyzed by the sandwich ELISA, the EPS in the com could be detected in the concentration range of $0.1-1.6{\;}\mu\textrm{g}/g$.

• Journal of Microbiology and Biotechnology
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• 제16권10호
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• pp.1613-1621
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• 2006

Isolation and identification of algal lytic bacteria were carried out. Nine strains of algal lytic bacteria were isolated by the double-layer method using Anabaena flos-aquae as a sole nutrient. The isolate, AFK-13, showing the highest algal lytic activity was identified as Sinorhizobium kostiense based on the l6S rDNA sequence. The algal lytic experiments of the culture supernatants of AFK-13 demonstrated that the bacterial cell growth reached a maximum at 36-h culture, but the supernatant of 72-h culture exhibited the highest activity. Components among the extracellular products in the crude enzyme of the supernatant from S. kostiense AFK-13 culture were responsible for degradation of cell walls of Anabaena flos-aquae. Algal lytic assay tests of the culture supernatants suggest that the main substances for algal lytic activity could be proteinaceous. The activity of glucosidase was observed highly by polysaccharolytic analysis using the crude enzyme from S. kostiense AFK-13, whereas activities of galactosidase, mannosidase, rhamnosidase, and arabinosidase were also detected in low levels. The molecular weights (MW) of ${\alpha}-\;and\;{\beta}$-glucosidases were estimated to be approximately 50-100 kDa by the ultrafiltration method.

• 한국미생물·생명공학회지
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• 제23권1호
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• pp.53-59
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• 1995

A protease isolated and purified 51 fold from the culture filtrate of a soil bacterium, Bacillus sp. KUN-17, which was appeared to be a monomeric protein with molecular weight of 38, 000 daltons, was suggested to be involved in the serine (-alkaline) protease (E.C 3.4.21.14) since its activity was selectively inhibited by phenylmethylsulfonyl fluoride (PMSF) and required 40$\circ$C and pH 10.5 for optimal condition. The half-life of the enzyme activity was 1 hr at 55$\circ$C, and the activity was maintained even under high concentrations of SDS or urea. The enzyme was indicated to perform random proteolysis from the fact that most of the chromogenic substrates employed were hydrolyzed by the enzyme. The affinity of the enzyme for natural proteins was approximately 10-times higher than ester compounds, and both substrates showed mutual inhibitory effect competitively for the enzyme activity.

• Journal of Microbiology
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• 제41권1호
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• pp.58-62
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• 2003

An extracellular pretense of Bacillus amyloliquefaciens S94 was purified to apparent homogeneity. The enzyme activity was strongly inhibited by general inhibitor for serine protease, PMSF, suggesting that the enzyme is a serine pretense. The purified enzyme activity was inhibited by leucine peptidase inhibitor, bestatin, suggesting that the enzyme is a leucine endopeptidase. The maximum proteolytic activity against different protein substrates occurred at pH 10, 45$^{\circ}C$ (protein substrate) and pH 8, 45$^{\circ}C$ (synthetic substrate). The purified enzyme was specific in that it readily hydrolyBed substrates with Leu or Lys residues at P$_1$ site. The pretense had characteristics of a cold-adapted protein, which was more active for the hydrolysis of synthetic substrate in the range of 15$^{\circ}C$ to 45$^{\circ}C$, specially at low temperature.

• 미생물학회지
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• 제38권1호
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• pp.50-53
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• 2002

백강균(Beauveria bassiana DGUM 34001)은 $24^{\circ}C$의 온도와pH 7.0의 초기 pH에서 최적의 균사체 생육을나타내었다. 사용한 복합배지 중 mushroom complex배지(MCM)에서 가장 우수한 균사생육을 나타내었다. Czapek-Dox 한천배지를 최소배지로 각각 탄소원, 질소원 및 인산원의 영향을 조사한 결과, glucose, soytone 및 sodium phosphate ($NaH_2$$PO_4$)에서 가장 우수한 균사 생육을 보여주었다. MCM액체배지에서 균사 배양 후 세포외 효소 활성을 측정한 결과,$\alpha$-amylase, lipase, chitinase, CMCase및 pretense의 비효소활성은 각각 297.0, 0.058, 0.33, 0.21 및 22.8 units/mg protein이었다. Casein과 soluble chitosan을 첨가할 경우 세포외 분비 pretense와 chitinase의 효소 활성 이 증가되었다.

• 미생물학회지
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• 제28권3호
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• pp.199-203
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• 1990

남극 토양생태계 세균의 분포와 세균의 생화학적 특성을 알아보기 위해 King George섬의 세종기지(한국)와 Te-niente Jubany기지(아르헨티나) 주변의 토양 17점을 남극 하계 기간(1989년 2월) 동안 채취하여 종속영양 세균의 수오 채외 효소활성도를 측정하였다. 종속영양 세균수는 시료채취지역과 배양온도가 다른 조건에서 자란 세균의 경우에도 그 평균값이 비숫한 경향을 보이고 있으나 protease는 $4^{\circ}C$에서 자란 세균의 경우 92%로서 특히 높게 나타났다. 배양온도에 따라 매우 변화가 켰으며 산술평균값은 배양온도 $37^{\circ}C$, $25^{\circ}C$, $4^{\circ}C$에서 각각 $2.5\times 10^{4}$, $2.7\times 10^{7}$, $6.9\times 10^{5}$ CFU/$cm^{3}$soil로 나타났다

• 미생물학회지
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• 제30권5호
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• pp.383-390
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• 1992

제주도를 중심으로 한 남해해역 ($123^{\circ}$/30'E-$128^{\circ}$/30'E, $32^{\circ}00N-34^{\circ}$ 30'N)을 대상으로 퇴적토 시료를 1988년 7월 31일부터 8월 10일, 1989년 3월 9일 부텨 3월 13일에 걸쳐 2회 채취하였다. 퇴적토시료는 숙성된 해수와 증류수를 각각 넣은 ZoBell 2216 E 평판배지를 사용하여 해양 세균과 담수 새균을 계수하였고 종속영양 세균분포와 세포외 효소활성을 측정 분석하였다. 종석영양세균의 분포는 하계의 경우 해양 세균과 담수 세균 각각 $0.3{\times}10^{6}~15{\times}10^{6}/cm^{3}$ sediment, $0.3{\times}10^{7}~30{\times}10^{7}/cm^{3}$ sediment, $0.01{\times}10^{6}~19{\times}10^{6}/cm^{3}$ sediment로 나타났다. 1989 년 춘계, sediment 표층 종속영양 세균의 절대값은 정점에 따라 1988 년 하계에 비해 최고 100배 이상으로 상승하였고, 대부분의 정점에서 많은 세균수를 관찰할 수 있었다. 한편 총세균에 대한 담수세균의 비율은 하계보다 춘계에 항승하는 결과를 나타냇다. 퇴적토 함수율은 하계보다 춘계에 낮게 나타났으나 유기물량을 나타내는 회분량은 높음을 알 수 있었다. 즉 춘계에는 하계에 비해 입자가 큰 최적토가 많이 분포되어 있으나 유기물의 유입이 많음을 시사한다. 이와 같은 결과를 종합해 볼때 남해해열 퇴적토 세균군집은 계절에 따라 매우 높은 분포를 보이고 이 요인으로는 육수의 유입으로 인한 영향을 들 수 있다. 또한 춘계에 해수의 혼합으로 무기영양물질 공급이 일어나고 이어 증식한 프랑크톤이 침강하여 퇴적토에 유입되므로써 세포외 효소 활성력이 높은 세균에 의해 고분자물질이 분해되고 이로 인한 영양물질 공급이 세균상승의 원인 중 하나일 것으로 사료된다.