|
|
|
| Cloning and Expression of the Linoleate Isomerase Gene from Lactobacillus plantarum ZS2058 in Kluyveromyces lactis GG799 |
| ZHU Jing-hua,CHEN Hai-qin,ZHANG Bai-xi,TIAN Feng-wei,ZHAO Jian-xin,CHEN Wei,ZHANG Hao |
| State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, Wuxi 214122, China |
|
|
|
Abstract Objective: To express the Linoleate isomerase gene of Lactobacillus plantarum in Kluyveromyces lactis. Method: Linoleate isomerase （LAI）gene was amplified by PCR from chromosome of Lactobacillus plantarum ZS2058, then the gene was cloned into Kluyveromyces lactis expression vector pKLAC1, the recombinant plasmid pKLAC1-LAI was transformed into Kluyveromyces lactis GG799 by electroporation, The expression level and the activity of recombinant enzyme were detected by SDS-PAGE and Gas Chromatogram . Result: It was demonstrated that a 67 kDa protein which was equal to LAI in molecular weight was secreted in supernatant culture . The analysis of gas chromatography showed that there was a noticeable peak in the retention time of 30.304 min in recombinant broth, the peak was Conjugated Linoleic Acid. Conclusion: It showed that the linoleate isomerase gene from L.plantarum ZS2058 had been expressed and secreted by Kluyveromyces lactis GG799 . Enzyme activity experiments showed that the recombinant enzyme can converse about 26% LA into CLA.
|
|
Received: 12 October 2009
Published: 29 April 2010
|
|
|
| ［1］ Peng S S, Deng M D. Purification and characterization of a membranebound linoleic acid isomerase from Clostridium sporogenes. Enzyme and Microbial Technology, 2007, 40(4): 831839.
［2］ 钮晓燕，陈卫，田丰伟等．植物乳杆菌ZS2058在磷酸盐缓冲液体系中生物转化共轭亚油酸．微生物学报，2007，47(2)： 244248. Niu X Y, Chen W, Tian F W, et al. Acta Microbiologica Sinica, 2007,47(2): 244248.
［3］ Rosson R A, Grund A D. Linoleate isomerase. World Patent,100846. 2001
［4］ Ellen H, Claudia K, Christian P, et al. Production of( 10E, 12Z) conjugated linoleic acid in yeast and tobacco seeds. Biochimoca et Biophysica Acta, 2005, 1738: 105 114.
［5］ Albert J J, van Ooyen1, Peter Dekker, et al. Heterologous protein production in the yeast Kluyveromyces lactis. FEMS Yeast Res, 2006，6:381392.
［6］ Jeremiah D Read, Paul A Colussi, Mehul B, et al. Acetamide selection of Kluyveromyces lactis cells transformed with an integrative vector leads to highfrequency formation of multicopy strains. Applied and Environmental Microbiology, 2007，8：50885096.
［7］ 许庆炎，陈海琴，田丰伟,等．植物乳杆菌ZS2058生物转化产物共轭亚油酸的分析方法的研究．食品与发酵工业 ，2008，34(1)：110114. Xu Q Y, Chen H Q, Tian F W, et al. Food and Fermentation Industries, 2008, 34(1): 110114.
［8］ Sambrook J, Fritsch E F, Maniatis T. Molecular Cloning: A Laboratory Manual. 2nded. New York: Cold Spring Harbor Laboratory Press,1989.
［9］ Thompson J R, Register E, Curotto J, et al. An improved protocol for the preparation of yeast cells for transformation by electroporation. Yeast, 1998, (14): 565571.
［10］ Colussi PA, Taron C H. Appl Environ Microbiol, 2005, 71: 70927098. |
|
Viewed |
|
|
|
Full text
|
|
|
|
|
Abstract
|
|
|
|
|
Cited |
|
|
|
|
| |
Shared |
|
|
|
|
| |
Discussed |
|
|
|
|
