• Journal of Microbiology and Biotechnology
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• 제24권10호
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• pp.1382-1388
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• 2014

Proteus vulgaris K80 lipase was expressed in Escherichia coli BL21 (DE3) cells and immobilized on amine-terminated magnetic microparticles (Mag-MPs). The immobilization yield and activity retention were 84.15% and 7.87%, respectively. A homology model of lipase K80 was constructed using P. mirabilis lipase as the template. Many lysine residues were located on the protein surface, remote from active sites. The biochemical characteristics of immobilized lipase K80 were compared with the soluble free form of lipase K80. The optimum temperature of K80-Mag-MPs was $60^{\circ}C$, which was $20^{\circ}C$ higher than that of the soluble form. K80-Mag-MPs also tended to be more stable than the soluble form at elevated temperatures and a broad range of pH. K80-Mag-MP maintained its stable form at up to $40^{\circ}C$ and in a pH range of 5.0-10.0, whereas soluble K80 maintained its activity up to $35^{\circ}C$ and pH 6.0-10.0. K80-Mag-MPs had broader substrate specificity compared with that of soluble K80. K80-Mag-MPs showed about 80% residual relative activity after five recovery trials. These results indicate the potential benefit of K80-Mag-MPs as a biocatalyst in various industries.

• KSBB Journal
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• 제20권5호
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• pp.338-340
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• 2005

변성제 (urea)와 환원제에 의해 완전히 풀린 상태의 lipase도 PEG에 의해 수식되는 것을 관찰하였다. 또한 mPEG-aldehyde로 수식된 mono-PEGylate과 di-PEGylate을 변성제와 환원제를 이용해 unfolding 시킨 후 희석에 의한 재접힘 시킨 결과, lipase에 공유결합된 PEG 분자는 재접힘 수율에 거의 영향을 미치지 않는 것으로 나타났다. 따라서 내포체 단백질을 대상으로 변성된 상태에서 PEGylation시킨 후 in vitro 재접힘 공정을 통해 PEGylation된 상태의 재생된 단백질을 회수할 수 있는 통합공정의 타당성을 제시하였다.

• Journal of Microbiology and Biotechnology
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• 제21권11호
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• pp.1159-1165
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• 2011

Biodiesel is methyl and ethyl esters of long-chain fatty acids produced from vegetable oils or animal fats. Lipase enzymes have occasionally been used for the production of this biofuel. Recently, biodiesel production using immobilized lipase has received increased attention. Through enhanced stability and reusability, immobilized lipase can contribute to the reduction of the costs inherent to biodiesel production. In this study, methanol-tolerant lipase M37 from Photobacterium lipolyticum was immobilized using the cross-linked enzyme aggregate (CLEA) method. Lipase M37 has a high lysine content (9.7%) in its protein sequence. Most lysine residues are located evenly over the surface of the protein, except for the lid structure region, which makes the CLEA preparation yield quite high (~93%). CLEA M37 evidences an optimal temperature of $30^{\circ}C$, and an optimal pH of 9-10. It was stable up to $50^{\circ}C$ and in a pH range of 4.0-11.0. Both soluble M37 and CLEA M37 were stable in the presence of high concentrations of methanol, ethanol, 1-propanol, and n-butanol. That is, their activities were maintained at solvent concentrations above 10% (v/v). CLEA M37 could produce biodiesel from olive oil and alcohols such as methanol and ethanol. Additionally, CLEA M37 generated biodiesel via both 2-step methanol feeding procedures. Considering its physical stability and reusability, CLEA M37 may potentially be used as a catalyst in organic synthesis, including the biodiesel production reaction.

• Fisheries and Aquatic Sciences
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• 제12권1호
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• pp.16-23
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• 2009

Lipids were extracted from marine rotifer, Brachionus rotundiformis in order to examine the functionality of lipid enzymatic modification. The fatty acids, palmitic, linoleic, oleic and stearic acids were the dominant forms accounting for approximately 35.8%, 21.5%, 15.9% and 7.7% of the total lipid content, respectively. Lipid fractions were categorized as neutral lipids (38.5%), glycolipids (45.9%) and phospholipids (17.6%), and after extraction from the rotifer were isolated by thin-layer chromatography (TLC) as free fatty acids (FFA), monoacylglycerol (MAG), diacylglycerol (DAG) and triacylglycerol (TAG). The production of polyunsaturated fatty acid (PUFA) concentrate from rotifer lipids was studied using lipase-catalyzed hydrolysis. In addition, rotifer lipids were modified by hydrolysis using lipases such as porcine pancreas, Candida rugosa and Rhizomucor miehei. The lipase from Rhizomucor miehei was effective in extracting linoleic acid (C 18:2), while the lipase from Candida rugosa was effective in palmitic acid (C16:0) extraction.

• International Journal of Industrial Entomology and Biomaterials
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• 제17권1호
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• pp.143-151
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• 2008

A muscle-specific lipase gene of the bumblebee Bombus ignitus was cloned and characterized. This gene, which we named Bi-Lipase, consists of seven exons encoding 317 amino acid residues. Bi-Lipase possesses all the features of lipases, including GXSXG consensus motif and Ser-Asp-His catalytic triad. Expressed as a 37-kDa polypeptide in baculovirus-infected insect Sf9 cells, recombinant Bi-Lipase showed an optimal pH of 9.0 and exhibited its highest catalytic activity at $40^{\circ}C$. Furthermore, through the addition of tunicamycin to the recombinant virus-infected Sf9 cells, recombinant Bi-Lipase was found to be N-glycosylated. Northern and western blot analyses indicated that Bi-Lipase was expressed in the wing, thorax, and leg muscles. These results show that Bi-Lipase is a muscle-specific lipase, suggesting a possible role of Bi-Lipase in the utilization of lipids for muscular activity in B. ignitus.

• 한국응용곤충학회지
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• 제46권1호
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• pp.131-139
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• 2007

곤충으로부터 유용 효소생산 미생물의 탐색 과정에서 우수한 lipase 생산균주 9종을 분리하고 lipase 생산능을 조사하였다 16S rDNA 분석 결과 분리된 균주는 주로 Serratia 속, Pseudomonas 속, Burkholderia 속에 속하는 그람음성균들로 분석되었다. 그 중 lipase 생산능이 가장 우수한 균주를 선별하고 16S rDNA 서열분석 및 생리 생화학적 분석 결과를 바탕으로 Burkholderia sp. HY-10으로 동정하였으며 균주의 lipase생산특성을 조사하였다. 이 균주는 톱하늘소의 장으로부터 분리되었으며 olive oil을 탄소원으로 포함하는 배지에서 배양하였을 때 세포밀도에 의존하여 lipase의 생산이 유도되는 특성을 나타내었고 0.5%의 yeast extract와 0.5%의 olive oil이 포함된 M9배지에서 $30^{\circ}C$, 36-42시간의 배양에 의해 lipase의 생산이 최대치에 도달하였다.

• Journal of Microbiology and Biotechnology
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• 제24권5호
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• pp.619-628
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• 2014

To identify lipase LipA (PFL_0617) from Pseudomonas protegens Pf-5, a lipA deletion mutant (Pf0617) and a complementary strain (Pf0617lipA) were constructed, and their effects on the lipase production were examined. Pf0617 remarkably decreased its whole-cell lipase activity, whereas Pf0617lipA made its whole-cell lipase activity not only restore to wild-type level but also get a further increment. However, the deletion and overexpression of lipA did not affect the extracellular lipase activity. In addition, the unbroken whole cells of these strains were able to catalyze the hydrolysis of membrane-permeable p-nitrophenyl esters, but could not hydrolyze the membrane-impermeable olive oil. These results confirmed that LipA was an intracellular lipase and Pf-5 could also be used as a natural whole-cell biocatalyst. To evaluate the potential of Pf-5 as a whole-cell biocatalyst and separately characterize the whole-cell LipA, the properties of the whole-cell lipases from Pf-5 and Top10lipA were characterized. The results demonstrated that both Pf-5 and Top10lipA exhibited high tolerance to alkaline condition, high temperature, heavy metal ions, surfactants, and organic solvents. Taken together, lipA can realize functional expression in E. coli Top10, and Pf-5 and Top10lipA as whole-cell biocatalysts may have enormous potential in applications.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 1986년도 추계학술대회
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• pp.523.2-523
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• 1986

The hydrolysis of olive oil was attempted with immobilized C. rugosa lipase in the reverse phase solvent system. (i.e. immobilized wet particles is dispersed in continuous phase olive oil or organic solvents containing olive oil). Sephadex LH-20 and LH-60 were used as the supports that can be used in organic solvents. The water content of wet particles of sephadex LH-20 and LH-60 were about 72% (w/w) and 85% (w/w), respectively Both swollen gels with 0.05M buffers adsorbed about 18% of lipase dissolved. They were easily dispersed in liquid olive oil or in organic solvents. The effects of organic solvents on the stability and catalytic activity of the lipase have been examined. The results revealed that isooctane is superior to the other solvents examined for enzymatic fat spliting in reverse phase system. Kinetics of enzymatic hydrolys of olive oil by immobilized lipase has been investigated in a batch reactor. Effects of pH and temperature on the lipase were studied. The substrate concentration was influenced positively on the thermal stability.

• Archives of Pharmacal Research
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• 제26권12호
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• pp.997-1001
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• 2003

This paper deals with chiral enzymatic resolution of 4-arylthio-2-butanols by lipase to prepare potential intermediates of $\beta$-lactam antibiotics. Among several lipases employed, lipase P type enzyme gave the highest ee value to prepare (R)-4-arylthio-2-butyl acetate. The enzymatic resolution of phenyl substituted alcohol (6a) using lipase P showed the highest ee value (99.7%) among those of 4-arylthio-2-butanol derivatives. Lipase P mediated hydrolysis of acylester 7a gave also (R)-alcohol 6a selectively. For determination of enantiomeric purity of these enzymatic resolved analytes, liquid chromatographic analysis was performed using two coupled Chiralcel OD and (R,R)-WhelkO chiral column.

• Bulletin of the Korean Chemical Society
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• 제24권9호
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• pp.1269-1275
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• 2003

The quaternary carbon-containing alcohols (1-6) were resolved enantioselectively by various lipases such as PFL (Pseudomonas fluorescens lipase), LAK (Pseudomonas fluorescens lipase), CRL (Candida rugosa lipase) and PCL (Pseudomonas cepacia lipase). The enzymatic resolution of racemic alcohol $({\pm})-2$ gave the excellent enantioselectivity in favor of (S)-2d in 99% ee, while those of the racemic alcohols (1, 3, 4, 5 and 6) gave the resolved alcohols with moderate to good enantioselectivity. Also, their absolute configurations were determined by chemical transformation to the known compounds.