重组大肠杆菌产Bacillus subtilis 168来源γ-谷氨酰转肽酶的摇瓶发酵优化

Fermentation Optimization of Recombinantγ-Glutamyltranspeptidase from Bacillus subtilis 168 in E. coli

为实现来源于Bacillus subtilis的γ-谷氨酰转肽酶（GGT）高效胞外分泌表达,以基因工程菌E.coli BL21（DE3）/p ET-20b（＋）/ggt为出发菌株,对其发酵产酶的诱导条件和培养基进行优化。最终确定其最优发酵培养基为：10 g/L甘油,21 g/L酵母粉,7 g/L蛋白胨,130 mmol/L磷酸盐;发酵条件为：37℃、200 r/min培养4 h,加入0.2 mmol/L IPTG后转入25℃,诱导40~48 h,优化后酶活达到81.2 U/m L,是未优化前的1.9倍。本研究结果为目前国内报道大肠杆菌摇瓶发酵产γ-谷氨酰转肽酶的最高酶活,为该酶的工业化生产奠定了基础。

In the present study,in order to improve the production of the recombinant Bacillus subtilis γ-glutamyltranspeptidase in E.coli BL21（DE3）/p ET-20b（＋）/ggt,the culture conditions and medium compositions were investigated and optimized in shake flasks. The optimal fermentation medium was as follows：10 g/L glycerol,21 g/L yeast extract,7 g/L peptone,and 130 mmol/L PO43＋.In addition,the optimal culture conditions were firstly cultured at 37 ℃,200 r/min for 4 h,and then induced by 0.2 mmol/L IPTG at temperature 25 ℃ and 40~48 h. Under these conditions,the maximal enzyme activity reached 81.2 U/m L,which was 1.9 times as high as that not optimized.