β-1,4-葡聚糖内切酶基因的克隆及在大肠杆菌中的表达

利用PCR方法从一株短小芽孢杆菌H9克隆了编码葡聚糖内切酶的基因（该序列已经被已收录于GeneBank，登录号为EF620915），序列分析表明该基因读码框为1980bp，编码659个氨基酸。预测的酶由两个不连续的结构域组成,其一为N-端催化结构域，由糖基水解酶家族9组成，第二个结构域为C-端底物结合结构域，由碳水化合物绑定结构域家族3组成。将基因构建在大肠杆菌表达载体pET20b中得到重组质粒pET20b-EglA,将重组载体转化至大肠菌株BL21(DE3)菌株,基因在大肠杆菌中得到了良好的分泌表达, SDS-电泳图谱表明酶的分子大小约为73KD。

cloning and expression of a endoglucanase gene in Escherichia.coli

The endoglucanase encoding gene was cloned from Bacillus pumilus H9 and the gene sequence is 1980bp ,coding 659 amino acids was found.. The predicted protein ,EglA ,consisted of an N-terminal catalytic domain belonging to glycoside hydrolase family 9 and a C-terminal cellulose-binding domain belonging to carbohydarte 3. The EglA gene was cloned into Escherichia.coli expression vector pET20b and the construct was transformed to a E.coli BL21(DE3) strain. EglA was Secreted expressed in Escherichia.coli and the molecular weight approximately is 73KD.