• 한국미생물·생명공학회지
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• 제48권4호
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• pp.480-490
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• 2020

A novel lipase gene, Lip-1420, was isolated from a metagenomic library constructed from reed marsh from Mt. Jumbong in Korea, comprising 112,500 members of recombinant plasmids. The DNA sequence of Lip-1420-subclone (5,513 bp) was found to contain at least 11 ORFs according to the GenBank database. The ORF-3 gene was inserted into the pET21a plasmid containing the C-terminal 6-His tag and transformed into E. coli BL21(DE3) to express the recombinant lipase protein. Lip-1420 was purified using a fast protein liquid chromatography system. The gene was registered in GenBank (MH628529). The values of Km and Vmax were determined as 0.268 mM and 1.821 units, respectively, at 40℃ and pH 8.0, using p-nitrophenyl palmitate as the substrate. This lipase belongs to family IV taxonomically because it has conserved HGGG and GDSAG motifs in the constitutive amino acid sequence. According to the predicted structural model, the binding sites are represented by residues H78, G81, D150, S151, A152, V181, and D236. Finally, Lip-1420 showed triolein selectivity for methanolysis between triolein (18:1) and tristearin (18:0) substrates. Further study of the selective mechanism and structure-function relationship of this new lipase could be useful for more practical applications.

• 한국미생물·생명공학회지
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• 제28권4호
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• pp.223-227
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• 2000

A novel two-step acetone treatment method was developed to enhance the enantioselectivity of Candida rugosa lipase (CRL) toward the hydrolysis of racemic naproxen methyl ester. The acetone-teated CRL was considerably more enantioselective than the crude CRL, yielding an enantiomeric excess of 98~100%. The crude and acetone-treated CRLs were subjected to anion exchange chromatography, and their chromatography profiles were compared. In consequence, both chromatography profiles were found to be almost identical, resulting in two separate lipase peaks (lipase A and B). The lipase B, which is known to be less enantioselective, was treated with acetone using a two-step treatment method. The enantioselectivity of acetone-treated lipase B was dramatically increased, yielding an enantiomeric excess of 99%.

• Journal of Microbiology and Biotechnology
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• 제12권1호
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• pp.142-144
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• 2002

Two different extracellular lipases were produced by an anaerobic bacterium, Selenomonas lipolytica. A major lipase, lipase I, was isolated, which showed optimum activity at pH 6.0 and at $45^{\circ}C$. It showed a molecular weight of 240 kDa and was a tetramer of a subunit having molecular weight of 60 kDa, which is different from the known bacterial lipases.

• Journal of Microbiology and Biotechnology
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• 제17권6호
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• pp.1054-1057
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• 2007

Lipases are industrially useful versatile enzymes that catalyze numerous different reactions including hydrolysis of triglycerides, transesterification, and chiral synthesis of esters under natural conditions. Although lipases from various sources have been widely used in industrial applications, such as in food, chemical, pharmaceutical, and detergent industries, there are still substantial current interests in developing new microbial lipases, specifically those functioning in abnormal conditions. We screened 17 lipase-producing yeast strains, which were prescreened for substrate specificity of lipase from more than 500 yeast strains from the Agricultural Research Service Culture Collection (Peoria, IL, U.S.A.), and selected Yarrowia lipolytica NRRL Y-2178 as a best lipase producer. This report presents new finding and optimal production of a novel extracellular alkaline lipase from Y. lipolytica NRRL Y-2178. Optimal culture conditions for lipase production by Y. lipolytica NRRL Y-2178 were 72 h incubation time, $27.5^{\circ}C$, pH 9.0. Glycerol and glucose were efficiently used as the most efficient carbon sources, and a combination of yeast extract and peptone was a good nitrogen source for lipase production by Y. lipolytica NRRL Y-2178. These results suggested that Y. lipolytica NRRL Y-2178 shows good industrial potential as a new alkaline lipase producer.

• Journal of Applied Biological Chemistry
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• 제50권2호
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• pp.46-51
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• 2007

Lipases are industrially useful versatile enzymes that catalyze numerous different reactions. Among lipases functioning under extreme conditions, alkaline lipase is useful in detergent industry. Lipase from yeast strain Yarrowia lipolytica NRRL Y-2178 was most active under alkaline condition, and initial medium pH for most lipase production was also alkaline [Lee et al., 2007, J Microbiol Biotechnol, 17(6)]. High lipase production was achieved using Y. lipolytica NRRL Y-2178. Optimal incubation time for lipase production at $25^{\circ}C$ was 72 h. Optimal temperature, when incubated for 72 h, was $27.5^{\circ}C$. Lipase production but not cell growth was very sensitive to concentrations of glucose and glycerol as efficient carbon sources, showing optimal concentrations of 1.0 and 1.5% (w/v), respectively. Lipase production was highly stimulated by $Ca^{2+},\;K^+,\;and\;Na^+$, but was inhibited by $Co^{2+},\;Cu^{2+},\;Mn^{2+},\;Na^+,\;and\;Fe^{2+}$. Maximum lipase production at 0.1 mM $Ca^{2+}$ for 72 h incubation at $27.5^{\circ}C$ was 649 units/mL.

• Journal of Applied Biological Chemistry
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• 제48권2호
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• pp.84-88
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• 2005

To develop functional food and anti-obesity drug through inhibition of dietary lipid absorption, inhibitory effects of herb extracts on pancreatic lipase were investigated. Due to high yield and simplicity of isolation, lipase inhibitor (ELI) was isolated from ethyl acetate extract of Eugenia aromaticum, which showed highest inhibitory activity, and characterized for development of novel functional material. Stability of ELI at high temperature and low pH was investigated. Results showed ELI is relatively stable under thermal and acidic conditions, reversible, and noncompetitive inhibitor of pancreatic lipase.

• Journal of Microbiology and Biotechnology
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• 제33권11호
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• pp.1531-1541
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• 2023

Lipase is a well-known and highly in-demand enzyme. During the last decade, several lipase optimization studies have been reported. However, production costs have always been a bottleneck for commercial-scale microbial enzyme production. This research aimed to optimize the conditions for lipase production by Limtongozyma siamensis DMKU-WBL1-3 via a One-Factor-At-a-Time (OFAT) approach combined with statistical methods while using a low-cost substrate. Results suggest that low-cost substrates can be substituted for all media components. An optimal medium was found, using response surface methodology (RSM) and central composite design (CCD), to consist of 0.50% (w/v) sweet whey, 0.40% (w/v) yeast extract (food grade), and 2.50% (v/v) palm oil with the medium pH adjusted to 4 under shaking flask cultivation. From an economic point of view, this work was successful in reducing production costs while increasing lipase productivity. The medium costs were reduced by 87.5% of the original cost while lipase activity was increased by nearly 6-fold. Moreover, lipase production was further studied in a 2-L stirred-tank fermentor. Its activity was 1,055.6 ± 0.0 U/ml when aeration and agitation rates were adjusted to 1 vvm and 170 rpm, respectively. Interestingly, under this optimal lipase production, the yeast showed accumulated lipids inside the cells. The primary fatty acid is a monounsaturated fatty acid (MUFA) that is typically linked to health benefits. This study hence reveals promising lipase production and lipid accumulation by L. siamensis DMKU-WBL1-3 that are worthy of further study.

• Journal of Microbiology and Biotechnology
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• 제30권7호
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• pp.1097-1103
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• 2020

Bacterial surface display systems have been developed for various applications in biotechnology and industry. Particularly, the discovery and design of anchoring motifs is highly important for the successful display of a target protein or peptide on the surface of bacteria. In this study, an efficient display system on Escherichia coli was developed using novel anchoring motifs designed from the E. coli mipA gene. Using the C-terminal fusion system of an industrial enzyme, Pseudomonas fluorescens lipase, six possible fusion sites, V¹⁴⁰, V¹⁷⁶, K¹⁷⁹, V²²⁶, V²³², and K²³⁴, which were truncated from the C-terminal end of the mipA gene (MV¹⁴⁰, MV¹⁷⁶, MV¹⁷⁹, MV²²⁶, MV²³², and MV²³⁴) were examined. The whole-cell lipase activities showed that MV¹⁴⁰ was the best among the six anchoring motifs. Furthermore, the lipase activity obtained using MV¹⁴⁰ as the anchoring motif was approximately 20-fold higher than that of the previous anchoring motifs FadL and OprF but slightly higher than that of YiaTR232. Western blotting and confocal microscopy further confirmed the localization of the fusion lipase displayed on the E. coli surface using the truncated MV¹⁴⁰. Additionally the MV¹⁴⁰ motif could be used for successfully displaying another industrial enzyme, α-amylase from Bacillus subtilis. These results showed that the fusion proteins using the MV¹⁴⁰ motif had notably high enzyme activities and did not exert any adverse effects on either cell growth or outer membrane integrity. Thus, this study shows that MipA can be used as a novel anchoring motif for more efficient bacterial surface display in the biotechnological and industrial fields.

• 한국식품영양학회지
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• 제32권1호
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• pp.27-32
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• 2019

The objective of this study was to evaluate novel usability as natural anti-obesity supplement of Selaginella tamariscina extract. The total phenol contents and total flavonoid contents were $60.29{\pm}3.11GAE\;mg/g$ and $14.90{\pm}0.34QE\;mg/g$, respectively. To evaluate anti-obesity activity of Selaginella tamariscina extract, pancreatic lipase inhibition activity as well as its inhibition effect of lipid accumulation in adipocytes were conducted by Oil Red O staining and lipolysis assay. The result of pancreatic lipase inhibition activity of S. tamariscina extract showed a wide range between 40 and 73% dose dependently. While the incubation of 3T3-L1 cells with S. tamariscina extract inhibited differentiation of preadipocytes and reduced lipid accumulation, the level of released free glycerol into culturing medium was increased in multiple concentrations. These results showed that S. tamariscina extract inhibit adipogenesis and pancreatic lipase activity. Thus, S. tamariscina extract can be a candidate for regulating lipid accumulation in obesity.

• 한국생명과학회:학술대회논문집
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• 한국생명과학회 2008년도 제49회 학술심포지움 및 국제학술대회
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• pp.51.1-51.1
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• 2008