• Journal of Plant Biotechnology
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• 제6권1호
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• pp.63-67
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• 2004

A simple procedure for the extraction of the lipolytic enzyme from rice bran has been developed. High activity of lipolytic enzyme was obtained by first defatting the rice bran to remove lipid components with various extraction conditions. Then, after rove cycles of aqueous extraction, rice bran lipolytic enzyme was purified using micro- and ultrafiltration apparatus. Lipolytic enzyme activity was estimated by its hydrolytic action of tributyrin. The result indicated that the standard activity curve of butyric acid showed that the potential rice bran enzyme is a hydrolytic lipase enzyme. In addition, it showed higher lipolytic activity and specific enzyme activity with further purification by micro- and ultrafiltration. The size of rice bran lipase enzyme was identified through 15 ％ SDS-PAGE. The molecular weight of the rice bran lipase enzyme was 41 kDa.

• 한국산학기술학회:학술대회논문집
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• 한국산학기술학회 2004년도 추계학술대회
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• pp.299-301
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• 2004

A simple procedure for the extraction of the lipolytic enzyme from rice bran has been developed. High activity of lipolytic enzyme was obtained by first defatting the rice bran to remove lipid components with various extraction conditions. Then, after five cycles of aqueous extraction, rice bran lipolytic enzyme was purified using micro- and ultrafiltration apparatus. Lipolytic enzyme activity was estimated by its hydrolytic action of tributyrin. The result indicated that the standard activity curve of butyric acid showed that the potential rice bran enzyme is a hydrolytic lipase enzyme. In addition, it showed higher lipolytic activity and specific enzyme activity with further purification by micro- and ultrafiltration.

• 생명과학회지
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• 제16권4호
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• pp.555-560
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• 2006

유용효소를 생산하는 균주를 가정하수로부터 분리하기 위하여 LBM, R2A, M9배지를 이용하여 다수의 균주를 분리하였다. 분리된 균주 중 1% tributyrin이 첨가된 배지에서 생육 활성대의 형성이 우수한 균주 1종을 최종적으로 선별하여 형태학적, 생리학적, 생화학적 특성을 관찰하였다. 16S rDNA 염기서열 분석결과 Acinetobacter Baumannii (99%)로 Acinetobacter 속에 속하는 균주임을 확인하고, Acinetobacter sp. BD5로 명명하였다. Acinetobacter sp. BD5는 $37^{\circ}C$와 $50^{\circ}C$에서 생육하는 것으로 보아 호열성 균주이며, 1% tributyrin과 oilve oil 첨가된 EL과 CE 고체배지와 Tween 20이 첨가된 LB 고체배지에서 생육활성대의 형성을 확인하여 이 균주가 lipolytic 효소를 생산하는 것으로 나타났다. 효소 활성의 최적 배양조건을 검토하고자 배양시간별 효소 활성을 측정한 결과, 배양 6시간 때인 대수 증식기에 가장 높은 효소활성을 나타내어 비교적 빠른 시간 내에 lipolytic 효소를 생성하는 것으로 나타났다. 또한 효소활성의 최적 온도는 $60^{\circ}C$로 $70-80^{\circ}C$에서 70%이상의 잔존활성을 보이는 것으로 나타나 Acinetobacter sp. BD5는 호열성 균주로 이 균주가 생산하는 lipolytic 효소도 내열성을 보이는 것으로 생각된다. 최적 pH는 9.0이며, pH 9.8-10.6범위에서 50% 이상의 활성이 유지되어 alkaline lipolytic 효소인 것으로 생각되어진다. Acinetobacter sp. BD5가 생산하는 lipolytic 효소는 비교적 넓은 pH 범위와 고온에서 활성이 유지되는 것으로 보아 폐유분해, 세제 합성과 유기물질 합성 등 생물공학분야와 산업적으로 잠재적 가치가 있을 것으로 생각된다.

• 생명과학회지
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• 제23권1호
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• pp.110-115
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• 2013

Extracellular 지질분해효소를 생산하는 균주 AR1은 일본 벳부 온천수에서 분리하였다. 분리된 균주의 16s rRNA 염기서열을 분석하고 계통학적으로 분류한 결과, AR1 균주는 신규 Geobacillus sp.에 속하는 것으로 동정되었다. 본 연구는 Geobacillus sp. AR1 균주의 extracellular 지질분해효소 생산을 향상시키기 위한 새로운 방법에 초점을 맞추었다. AR1 균주는 $35{\sim}75^{\circ}C$의 넓은 온도 범위에서 생육하였고 최적온도는 $65^{\circ}C$이었다. 생육을 위한 최적 pH는 6.5인 반면, 효소 생산을 위한 pH는 8.5로 차이점을 보였다. 배양 중에 지질 화합물의 첨가는 지질분해효소 생산을 유도하였고, soybean oil을 대수증식기에 첨가 했을 때 가장 효율적인 유도 효과를 나타내었다. 한편, 계면활성제는 지질분해효소의 생산을 유도하고 세포 내외의 위치에 영향을 줄 수 있다. AR1 균주는 정지기에 Tween 20을 첨가할 경우, 효소의 세포 외 분비 효율이 크게 증가하였다. 이들 결과를 바탕으로 soybean oil과 Tween 20을 각각 대수증식기와 정지기에 첨가함에 따라 extracellular 효소 생산이 대조구에 비해 2.4배 증가하는 것으로 확인 되었다.

• 생명과학회지
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• 제20권5호
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• pp.662-669
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• 2010

폐식용유에서 지방분해효소를 생산하는 세균을 분리하였고, PIBWIN 세균동정 방법으로 생리 생화학적 특성을 조사하여 확인한 결과 Pseudomonas sp. OME로 동정하였다. 여러 기질로 지방분해효소 생산을 조사한 결과 올리브유에서 6.1 U/ml의 생산력을 나타내었다. 물리적 인자인 배양시간, 온도. pH 및 올리브유와 효모 추출액의 영양인자에 의한 지방분해효소 생산 조건을 조사 하였다. 효소의 분비는 배양시간. 올리브유 와 효모 추출액의 농도에 강한 영향을 받았으며, RSM을 이용한 최적화는 이들 인자를 가지고 조사하였다. RSM을 이용한 지방분해효소 생산은 배양시간. 올리브유와 효모 추출액의 농도가 48 hr, 0.3 g, 및 0.9 ml에서 최적 생산조건을 나타냈다.

• Journal of Microbiology and Biotechnology
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• 제17권10호
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• pp.1655-1660
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• 2007

A metagenome is a unique resource to search for novel microbial enzymes from the unculturable microorganisms in soil. A forest soil metagenomic library using a fosmid and soil microbial DNA from Gwangneung forest, Korea, was constructed in Escherichia coli and screened to select lipolytic genes. A total of seven unique lipolytic clones were selected by screening of the 31,000-member forest soil metagenome library based on tributyrin hydrolysis. The ORFs for lipolytic activity were subcloned in a high copy number plasmid by screening the secondary shortgun libraries from the seven clones. Since the lipolytic enzymes were well secreted in E. coli into the culture broth, the lipolytic activity of the subclones was confirmed by the hydrolysis of p-nitrophenyl butyrate using culture supernatant. Deduced amino acid sequence analysis of the identified ORFs for lipolytic activity revealed that 4 genes encode hormone-sensitive lipase (HSL) in lipase family IV. Phylogenetic analysis indicated that 4 proteins were clustered with HSL in the database and other metagenomic HSLs. The other 2 genes and 1 gene encode non-heme peroxidase-like enzymes of lipase family V and a GDSL family esterase/lipase in family II, respectively. The gene for the GDSL enzyme is the first description of the enzyme from metagenomic screening.

• Journal of Microbiology and Biotechnology
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• 제15권5호
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• pp.1067-1072
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• 2005

A metagenomic library was constructed using a fosmid vector, and total genomic DNA was extracted directly from soil at Cisolok (hot spring area, Indonesia). This library was composed of 10,214 clones and screened for lipolytic enzyme on tributyrin agar plates. An esterase gene (estMa) was subcloned and sequenced from a positive lipolytic active clone. Esterase EstMa was encoded by a 954-bp open reading frame and showed low ($11-33\%$) amino acid similarity to known esterases. The amino acid sequence analysis demonstrated that the enzyme is a new member of lipolytic enzyme family VI. The estMa gene encodes a preprotein of 317 amino acids with a predicted molecular mass of 34,799 Da. The purified enzyme exhibited optimal activity at $50^{\circ}C$ and pH 6.5. The $K_m,\;and\;V_{max}$ values of EstMa for the hydrolysis of p-nitrophenyl valerate were $45.3\;{\mu}M$ and 4.45 U/mg, respectively.

• 한국염색가공학회:학술대회논문집
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• 한국염색가공학회 2008년도 제39차 학술발표회
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• pp.29-30
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• 2008

• Mycobiology
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• 제41권2호
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• pp.67-72
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• 2013

Pathogenic microbes secrete various enzymes with lipolytic activities to facilitate their survival within the host. Lipolytic enzymes include extracellular lipases and phospholipases, and several lines of evidence have suggested that these enzymes contribute to the virulence of pathogenic fungi. Candida albicans and Cryptococcus neoformans are the most commonly isolated human fungal pathogens, and several biochemical and molecular approaches have identified their extracellular lipolytic enzymes. The role of lipases and phospholipases in the virulence of C. albicans has been extensively studied, and these enzymes have been shown to contribute to C. albicans morphological transition, colonization, cytotoxicity, and penetration to the host. While not much is known about the lipases in C. neoformans, the roles of phospholipases in the dissemination of fungal cells in the host and in signaling pathways have been described. Lipolytic enzymes may also influence the survival of the lipophilic cutaneous pathogenic yeast Malassezia species within the host, and an unusually high number of lipase-coding genes may complement the lipid dependency of this fungus. This review briefly describes the current understanding of the lipolytic enzymes in major human fungal pathogens, namely C. albicans, C. neoformans, and Malassezia spp.

• 미생물학회지
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• 제30권3호
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• pp.171-186
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• 1992

Lipolytic characteristics of candia cylindracea lipase was studied by various triacylglycerols with different fatty acyl chains as substrate. The substrate was emulsified with gum arabic and the rate of hydrolysis was determined by pH stat method. The effects of gum concentration, pH, temperature, and $Ca^{2+}$ ion on the enzyme activities were examined. The results show that the effect of these factors are markedly depending on the structurla nature of substrates. The triolein was the best substrate among tested. Present study demonstrates that for characterization of lipolytic enzymes, it is critically important to select proper substrate and activator.r.