阿魏酸酯酶基因工程菌发酵条件优化

Optimization on fermentation condition of genetic engineering bacteria of feruloyl esterase

以本实验室前期构建的重组大肠杆菌工程菌株为基础,为提高阿魏酸酯酶的表达量,通过试验设计,对工程菌株目的蛋白可溶表达条件进行优化。实验采用250 mL三角瓶中,内含50 mL(Amp 100 mg/L)的LB培养基,分别研究诱导温度、诱导物IPTG浓度、诱导时机和诱导时间等对蛋白可溶表达量的影响。确定了阿魏酸酯酶工程菌可溶性表达最优条件为:过夜培养菌体以1∶100比例接种新鲜培养基,摇床转速200 rmp/min,在37℃下培养至OD600值为1.0时,添加终浓度为0.01 mM的IPTG进行诱导,诱导温度30℃,诱导时长为9 h,最终单位发酵液的酶活力为400 U/L。以上数据为阿魏酸酯酶重组工程菌的工业应用奠定了基础。

On the basis of the recombinant E. coli strains which were built in our laboratory,in order to improve the expression level of feruloyl esterase,the conditions of soluble protein expression were optimized through experimental design. Using 250 mL Erlenmeyer flask containing 50 mL（ Amp 100 mg /L）medium,induction temperature,inducer concentration,induced expression of soluble protein timing and induction time were studied. Identified feruloyl esterase engineered bacteria optimal conditions for soluble expression were incubated at 37 ℃ to OD600 value of 1. 0,adding a final concentration of 0. 01mM of IPTG,induction temperature 30 ℃,shaking speed 200 rmp /min,inoculum of 1%,while the induction time 9 h. After the fermentation 400 U /L enzyme activity was obtained. The above data make a basis for the recombinant engineering bacteria pilot fermentation.