Expression of Bacillus circullans Lactase Gene in E. coli and Properties of Expressed Lactase
Wang Yuanhuo1 Yao Bin1** Yuan Tiezheng1 Cao Shishu1 Zhang Wei2 Wang Yaru1 Fan Yunliu2
(1. Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China; 2 .Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China)
Abstract：Abstract : The lactase gene from Bacillus circulans was overexpressed in Escherichia coli and the expressed lactase had normal bioactivity. The properties of purified lactase expressed in E. coli and produced by Bacillus circulans were determined. The results revealed, between the lactase expressed in E. coli and produced by Bacillus circulans, that there had notable difference in enzyme properties in optimal pH value and pH stability, optimal temperature and temperature stability, Km value. The optimal pH value of the lactase expressed in E. coli was 5.0, whereas that of origin lactase produced by Bacillus circulans was 6.5. The optimal temperature of expressed lactase was 37 ℃ and 55 ℃, whereas that of origin lactase was 55 ℃. After treated for 2 h under pH 4.0, the remaining enzyme activity of expressed lactase was 85%, whereas that of origin lactase was only 5%. The thermal stability of expressed lactase was better than that of origin lactase, after treated for 10 minutes under 60 ℃, the remaining enzyme activity of expressed lactase was 30% , whereas origin lactase almost lost all of its activity. The expressed lactase had better metal ion resistance to Cu2+, Mn2+ and Fe2+, the remaining enzyme activity of expressed lactase and origin lactase in reaction system with 1mmol/L Fe2+ was 96% and 22%, respectively. The Km and Vmax values of expressed lactase had 285-fold decrease and 5.4-fold increase, respectively, compared to those of origin lactase. The lactase was overexpressed in E. coli, the expression level of lactase was high up to 33.102 U/mL. The properties of the lactase expressed in E. coli including high activity in pH 3.5～8.5, good thermal stability and metal ion resistance demonstrate that the enzyme is a good candidate in dairy industry.
王元火1 姚 斌1** 袁铁铮1 操时树1 张 伟2 王亚茹1 范云六2. 环状芽孢杆菌乳糖酶基因在大肠杆菌中的表达及酶学性质分析[J]. , 2003, 11(1): 83-88.
Wang Yuanhuo1 Yao Bin1** Yuan Tiezheng1 Cao Shishu1 Zhang Wei2 Wang Yaru1 Fan Yunliu2. Expression of Bacillus circullans Lactase Gene in E. coli and Properties of Expressed Lactase. , 2003, 11(1): 83-88.
