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| Recombinant Expression and Fermentation Optimization of Dictyoglomus thermophilum Cellobiose 2-Epimerase in Bacillus subtilis |
Xin-miao WANG,Kang ZHANG,Sheng CHEN,Jing WU( ) |
| 1 School of Biotechnology and Key Laboratory of Industrial Biotechnology Ministry of Education, International Joint Laboratory on Food Safety, State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China |
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Abstract A cellobiose 2-epimerase gene ce derived from Dictyoglomus thermophilum was synthesized and introduced into the expression vector pBSuL3 to construct a recombinant plasmid pBSuL3-ce and then transformed it into Bacillus subtilis. The intracellular activity of cellobiose epimerase reached 7.5U/ml after cultivated in TB culture for 48h. The enzymatic properties showed that the optimum pH of the enzyme was 8.5;the optimum temperature was 85℃, and the half-life of 85℃was 120min. In order to reduce the cost of fermentation, the fermentation medium was optimized.When 35g/L soybean meal was used as nitrogen source and 5g/L glycerin was used as carbon source, the intracellular activity reached 12.3U/ml. Then, according to the optimized condition of the shake flask, the culture was expanded in the 3L fermenter, and the intracellular activity of cellobiose epimerase reached 56U/ml, which was 8-fold higher than that of the shake flask culture.The lactulose was prepared by using the enzyme obtained by fermentation. When the lactose concentration was 400g/L, the reaction temperature was 85℃, the initial pH was 8.5, the enzyme amount was 20U/ml, the enzyme converted lactose to 51% lactulose.
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Received: 29 December 2018
Published: 05 August 2019
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Corresponding Authors:
Jing WU
E-mail: jingwu@jiangnan.edu.cn
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