Screening and Identification of High-yield Thermostable Lipase Producing Microorganisms

doi:10.13560/j.cnki.biotech.bull.1985.2015.01.022

Abstract

Abstract: Two strains of microorganisms（J2 and J3）with a high-yield lipase producing ability had been screened from dumplings’ steamer cushions, and identified as two variants of Aureobasidium sp. according to morphological observation and 26S rRNA gene（26S rDNA）sequencing. After being incubated at 30℃, 200 r/min for 3-5 d, the lipase activities of their fermentation supernatant were 10.61 U/mL and 14.43 U/mL, respectively, measured by spectrophotometry with p-nitrophenol palmitate（p-NPP）as a substrate. The investigation of physicochemical properties of the produced lipases（Lip2 and Lip3）showed that the optimum catalyzing temperature of Lip2 is 50℃ and there is no significant activity loss after being incubated at 50℃ for 5 h, while the optimum catalyzing temperature of Lip3 is 60℃ and there is no activity loss after being incubated at 40℃ for 5 h, but only 42.19% of lipase activity remained when incubated at 50℃ for 5 h. The results indicated that the lipases produced by strain J2 and J3 have good thermal stability and high optimal reaction temperature.

Key words: lipase, thermostability, Aureobasidium sp., 26S rDNA, growth curve

Li Yang, Cai Haiying, Zhao Minjie, Zhang Hui, Feng Fengqin. Screening and Identification of High-yield Thermostable Lipase Producing Microorganisms[J]. Biotechnology Bulletin, 2015, 31(1): 144-150.

References

[1] Tamalampudi S, Talukder MR, Hama S, et al. Development of recombinant Aspergillus oryzae whole-cell biocatalyst expressing lipase-encoding gene from Candida antarctica[J]. Appl Microbiol Biotechnol, 2007, 75（2）：387-395.
[2] Dizge N, Aydiner C, Imer DY, et al. Biodiesel production from sunflower, soybean, and waste cooking oils by transesterification using lipase immobilized onto a novel microporous polymer[J]. Bioresource Technology, 2009, 100（6）：1983-1991.
[3] 韦伟, 冯凤琴. Sn-1, 3位专一性脂肪酶在食品中的应用[J].中国粮油学报, 2012, 27（2）：122-128.
[4] 王海燕, 李富伟, 高秀华.脂肪酶的研究进展及其在饲料中的应用[J].饲料工业, 2007, 28（6）：14-17.
[5] Levisson M, van der Oost J, Kengen SW. Carboxylic ester hydrolases from hyperthermophiles[J]. Extremophiles, 2009, 13（4）：567-581.
[6] Esteban L, Jiménez MJ, Hita E, et al. Production of structured tria-cylglycerols rich in palmitic acid at sn-2 position andoleic acid at sn-1, 3 positions as human milk fat substitutes by enzymatic acidoly-sis[J]. Biochemical Engineering Journal, 2001, 54（1）：62-69.
[7] 刘燕雅, 黄平, 司圣乾, 等. P197E 与ep8 叠加突变对扩展青霉脂肪酶热稳定性的影响[J]. 微生物学通报, 2010, 37（2）：161-165.
[8] Mu?oz PA, Correa-Llantén DN, Blamey JM. Production, purification and partial characterization of four lipases from a thermophile isolated from deception island[J]. Lipids, 2013, 48（5）：527-533.
[9] Winkler UK, Stuckmann M. Glycogen, hyaluronate, and some other polysaccharides greatly enhance the formation of exolipase by Serratia marcescens[J]. Journal of Bacterology, 1979, 138（3）：663-670.
[10] Najjar A, Robert S, Guérin C, et al. Quantitative study of lipase secretion, extracellular lipolysis, and lipid storage in the yeast Yarrowia lipolytica grownin the presence of olive oil：analogies with lipolysis in humans[J]. Applied Microbiology and Biotechnology, 2011, 89（6）：1947-1962.
[11] 李俊峰, 李红芳, 段效辉, 等.耐有机溶剂脂肪酶产生菌的筛选及其粗酶酶学性质[J].食品科学, 2012, 33（3）：116-120.
[12] Simon L, Caye-Vaugien C, Bouchonneau M. Relation between pullulan production, morphological state and growth conditions in Aureobasidium pullulans：new observations[J]. Journal of General Microbiology, 1993, 139：979-985.
[13] 谷才恩.短梗茁霉胞外多糖的研究Ⅰ菌株的筛选[J].微生物学通报, 1985, 2：67-69.
[14] 周文化, 钟秋平.出芽短梗霉菌的微生物发酵及在食品工业中的应用[J].食品科学, 2004, 25：179-186.
[15] Liu Z, Chi Z, Wang L, et al. Production, purification and charac-terization of an extracellularlipase from Aureobasidium pullulans HN2.3 with potentialapplication for the hydrolysis of edible oils[J]. Biochemical Engineering Journal, 2008, 40（2）：445-451.
[16] Goldbeck R, Maugeri F, Goldbeck R. Screening, characterization, and biocatalytic capacity of lipases producing wild yeasts from Brazil biomes[J]. Food Science and Biotechnology, 2013, 22（1）：79-97.
[17] Mhetras NC, Bastawde KB, Gokhale DV. Purification and charac-terization of acidic lipase from Aspergillus niger NCIM 1207[J]. Bioresource Technology, 2009, 100（3）：1486-1490.
[18] Liu Z, Li X, Chi Z, et al. Cloning, characterization and expression of the extra cellular lipase gene from Aureobasidium pullulans HN2-3 isolated from sea saltern[J]. Antonie van Leeuwenhoek, 2008, 94（2）：245-255.