|Table of Contents|

Secretory expression of bile salt hydrolase gene from Lactobacillus plantarum BBE7 in Pichia pastoris(PDF)

Chinese Journal of Applied & Environmental Biology[ISSN:1006-687X/CN:51-1482/Q]

Issue:
2014 01
Page:
107-111
Research Field:
Articles
Publishing date:

Info

Title:
Secretory expression of bile salt hydrolase gene from Lactobacillus plantarum BBE7 in Pichia pastoris
Author(s):
DONG Zixing;  ZHANG Juan;  LEE Byonghoon;  DU Guocheng;  CHEN Jian;  LI Huazhong
1 School of Biotechnology, Jiangnan University, Wuxi 214122, China 2 Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, China 3 National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China 4 Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, China
Keywords:
Lactobacillus plantarum BBE7;  bile salt hydrolase;  Pichia pastoris;  secretory expression;  orthogonal experimental design
CLC:
Q936 : Q786
PACS:
DOI:
10.3724/SP.J.1145.2014.00107
DocumentCode:

Abstract:
When bile salt hydrolase (bsh) genes from Lactobacillus plantarum are expressed intracellularly and extracellularly by Escherichia coli, they usually form inclusion bodies. Therefore, Pichia pastoris, an efficient expression system, was used to achieve the secretory expression of bsh. Firstly, signal peptide α-factor was ligated with bsh by fusion PCR, and the fragment obtained was then cloned into vector pPIC9K, resulting in a recombinant plasmid pPIC9K-α-factor-BSH. Finally, this plasmid was linearized by Sal I and transformed into Pichia pastoris GS115. As analyzed by SDS-PAGE and BSH assay, the molecular weight and extracellular activity of recombinant BSH was determined to be 37.0 × 103 and 1.08 U mL-1, respectively. After optimization of fermentation conditions using orthogonal experimental design (L9 (34))，the extracellular BSH activity was 2.69 U mL-1, 1.49 times higher than before. This lays a solid foundation for purification, characterization and large-scale production of BSH.

References

1 Sridevi N, Vishwe P, Prabhune A. Hypocholesteremic effect of bile salt hydrolase from Lactobacillus buchneri ATCC 4005 [J]. Food Res Int, 2009, 42 (4): 516-520
2 牛治霞, 刘恩梅, 李晓东, 张柏林. 益生菌中胆盐水解酶(BSHs)研究进展[J]. 中国乳品工业, 2007, 35 (9): 35-40 [Niu ZX, Liu EM, Li XD, Zhang BL. Study progress of the bile salt hydrolases enzymes (BSHs) in vivo [J]. China Dairy Ind, 2007, 35 (9): 35-40]
3 Jiang J, Hang X, Zhang M, Liu X, Li D, Yang H. Diversity of bile salt hydrolase activities in different Lactobacilli toward human bile salts [J]. Ann Microbiol, 2010, 60 (1): 81-88
4 Begley M, Hill C, Gahan CG. Bile salt hydrolase activity in probiotics [J]. Appl Environ Microbiol, 2006, 72 (3): 1729-38
5 Hatakka K, Mutanen M, Holma R, Saxelin M, Korpela R. Lactobacillus rhamnosus LC705 together with Propionibacterium freudenreichii ssp shermanii JS administered in capsules is ineffective in lowering serum lipids [J]. J Am Coll Nutr, 2008, 27 (4): 441-447
6 黄茜, 黄璐, 潘道东, 杨瑶. 植物乳杆菌Lactobacillu plantarum Y1菌株胆盐水解酶基因(bsh)的克隆及重组表达[J]. 南京师大学报: 自然科学版, 2010, 33 (3): 91-96 [Huang Q, Huang L, Pan DD, Yang Y. Molecular cloning and expession of the bile salt gene (bsh) from Lactobacillus plantarum Y1 [J]. J Nanjing Norm Univ (Nat Sci Ed), 2010, 33 (3): 91-96]
7 战媛媛, 王长远, 于长青. 植物乳杆菌中胆盐水解酶基因的原核表达及纯化[J]. 中国生物制品学杂志, 2009, 22 (9): 876-879 [Zhan YY, Wang CY, Yu CQ. Prokaryotic expression of bile salt hydrolase gene of Lactobacillus plantarum and purification of expressed product [J]. Chin J Biologicals, 2009, 22 (9): 876-879]
8 Dong Z, Zhang J, Lee BH, Li H, Du G, Chen J. Secretory expression and characterization of a bile salt hydrolase from Lactobacillus plantarum in Escherichia coli [J]. J Mol Catal B-Enzym, 2013, 93: 57-64
9 Teng D, Fan Y, Yang YL, Tian ZG, Luo J, Wang JH. Codon optimization of Bacillus licheniformis β-1, 3-1, 4-glucanase gene and its expression in Pichia pastoris [J]. Appl Microbiol Biotechnol, 2007, 74 (5): 1074-1083
10 Dong Z, Zhang J, Lee B, Li H, Du G, Chen J. A bile salt hydrolase gene of Lactobacillus plantarum BBE7 with high cholesterol-removing activity [J]. Eur Food Res Technol, 2012, 235 (3): 419-427
11 Jana S, Deb J. Strategies for efficient production of heterologous proteins in Escherichia coli [J]. Appl Microbiol Biotechnol, 2005, 67 (3): 289-298
12 黄茜, 黄璐, 潘道东, 杨瑶. 重组融合蛋白MBP-BSH在大肠杆菌中的表达及其纯化, 功能鉴定[J]. 食品科学, 2012, 33 (7): 198-203 [Huang Q, Huang L, Pan DD, Yang Y. Expression in Escherichia coli, purification and functional charaterization of recombinant fusion protein MBP-BSH [J]. Food Sci, 2012, 33 (7): 198-203]
13 于长青, 战媛媛, 王长远. 胆盐水解酶基因在毕赤酵母中的表达[J]. 中国生物制品学杂志, 2010, 23 (9): 949-952 [Yu CQ, Zhan YY, Wang CY. Expression of bile salt hydrolase gene in Pichia pastoris [J]. Chin J Biol, 2010, 23 (9): 949-952]
14 耿宏伟, 侯红燕, 王丕武, 张建峰. pGAP-毕赤酵母表达系统的研究进展[J]. 农业机械, 2011 (20): 159-162
15 Kim GB, Miyamoto CM, Meighen EA, Lee BH. Cloning and characterization of the bile salt hydrolase genes (bsh) from Bifidobacterium bifidum strains [J]. Appl Environ Microbiol, 2004, 70 (9): 5603-12
16 Southgate V, Steyn A, Pretorius I, Van Vuuren H. Expression and secretion of Bacillus amyloliquefaciens alpha-amylase by using the yeast pheromone alpha-factor promoter and leader sequence in Saccharomyces cerevisiae [J]. Appl Environ Microbiol, 1993, 59 (4): 1253-1258
17 Zhang WY, Wu RN, Sun ZH, Sun TS, Meng H, Zhang HP. Molecular cloning and characterization of bile salt hydrolase in Lactobacillus casei Zhang [J]. Ann Microbiol, 2009, 59 (4): 721-726
18 Kim GB, Lee BH. Genetic analysis of a bile salt hydrolase in Bifidobacterium animalis subsp. lactis KL612 [J]. J Appl Microbiol, 2008, 105 (3): 778-90

Memo

Memo:
-
Last Update: 2014-03-06