Expression of Bile Salt Hydrolase Gene from Lactobacillus plantarum DPP8 in Escherichia coli and Its Enzymatic Properties

DING Ke; DUAN Jin; YU Zuhua; LI Wang; LI Yuanxiao; HE Wanling; ZHANG Chunjie; DING Panpan; WANG Yuqin; LIU Ning

doi:10.3969/j.issn.1006-267x.2020.07.043

2020 , Vol. 32 >Issue 7: 3333 - 3342

DOI: https://doi.org/10.3969/j.issn.1006-267x.2020.07.043

Expression of Bile Salt Hydrolase Gene from Lactobacillus plantarum DPP8 in Escherichia coli and Its Enzymatic Properties

DING Ke ,
DUAN Jin ,
YU Zuhua ,
LI Wang ,
LI Yuanxiao ,
HE Wanling ,
ZHANG Chunjie ,
DING Panpan ,
WANG Yuqin ,
LIU Ning

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1. Hongxiang Biological Feed Laboratory, Henan University of Science and Technology, Luoyang 471000, China;
2. Key Laboratory of Animal Disease and Public Health of Henan Province, Luoyang 471000, China;
3. Research Center of Breeding Engineering Technology for Meat Sheep of Henan Province, Luoyang 471000, China

Received date: 2020-01-28

Online published: 2020-07-15

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Abstract

The aim of this experiment was to clone the bile salt hydrolase (BSH) gene from Lactobacillus plantarum DPP8 and express it in Escherichia coli (E. coli) expression system, and to study its enzymatic properties. The BSH gene was cloned by PCR technique, and was inserted into vector pET32a(+) to construct the recombinant expression vector pET32a(+)-BSH, which was transformed into E.coli BL21. The recombinant strain was induced to express BSH by isopropylthiogalactosidehe (IPTG). The expressed BSH activity was detected by calibration method of taurine standard solution, and the enzymatic properties of the expressed BSH was studied. The results showed that a whole 975 bp sequence was obtained, and was also proved successfully that it was the BSH gene by homology comparison and phylogenetic tree analysis. Sodiumdodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) assay indicated that the colecular weight of the recombinant protein was approximately 56 ku, which was consistent with the expected size. The highest BSH activity was 22.81 U/mL in recombinant strain with the conditions at 37℃, 0.1 mmol/L IPTG induction 6 h. The reaction optimum temperature of the purified BSH was 37℃ and the enzyme can still maintain the good thermal stability under 40℃ for 1 h. The reaction optimum pH of the purified BSH was 6.5 and more than 90% enzyme activity was maintained at pH 3.0 to 9.0, 37℃ for 1 h. Mg²⁺, Al³⁺ and Fe³⁺ had strong inhibition effects on the BSH activity, but Na⁺, K⁺, Fe²⁺, Cu²⁺, Mn²⁺, Zn²⁺ and Ca²⁺ had no obvious effects on it. In conclusion, the BSH from Lactobacillus plantarum DPP8 is expressed in E. coli, successfully, the highest BSH activity is 22.81 U/mL, and it can keep high enzyme activity under 40℃ and within pH 3.0 to 9.0, but can be strong inhibited by Mg²⁺, Al³⁺ and Fe³⁺.

Key words： Lactobacillus plantarum; bile salt hydrolase; clone; expression; enzymatic properties

Cite this article

DING Ke , DUAN Jin , YU Zuhua , LI Wang , LI Yuanxiao , HE Wanling , ZHANG Chunjie , DING Panpan , WANG Yuqin , LIU Ning . Expression of Bile Salt Hydrolase Gene from Lactobacillus plantarum DPP8 in Escherichia coli and Its Enzymatic Properties[J]. Chinese Journal of Animal Nutrition, 2020 , 32(7) : 3333 -3342 . DOI: 10.3969/j.issn.1006-267x.2020.07.043

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