GST-SUMO-MT融合蛋白在大肠杆菌中的自诱导表达

Auto-inducing Expression of GST-SUMO-MT Fusion Protein in Escherichia coli

旨在考察在大肠杆菌中自诱导表达GST-SUMO-MT融合蛋白的可行性,并对自诱导培养条件及培养基成分进行优化,以提高蛋白产量。采用摇瓶培养,利用单因素和正交实验对工程菌自诱导的培养条件(诱导时间、诱导温度)和培养基组分(蛋白胨、酵母粉、甘油、葡萄糖、乳糖)进行优化,检测菌体浓度、可溶性目的蛋白的表达量。结果表明,自诱导培养基碳氮源的最优配比为:2%蛋白胨、2%酵母粉、0.3%甘油、0.05%葡萄糖和0.3%乳糖。重组菌自诱导表达最优发酵条件为:采用两阶段温度控制,37℃培养3 h,25℃继续培养13 h。此时可溶性目的蛋白的表达量和菌体浓度分别为LB培养基(IPTG诱导)的2.2倍和2.3倍。

The auto-inducing expression of GST-SUMO-MT fusion protein in Escherichia coli BL21（DE3）was studied. In order to increase the protein yield, we optimized both the auto-induction fermentation conditions and medium. Recombinant E. coli was cultured in shake flask, of which the culture condition（time and temperature for induction）and medium composition（tryptone, yeast extract, glycerol, glucose and lactose）were optimized by single factor and orthogonal tests, and bacterial concentration, expression level of soluble target protein were determined. Result showed that the optimal carbon and nitrogen composition ratio in auto induction medium was obtained as followed ：2% tryptone, 2% yeast extract, 0.3% glycerol, 0.05% glucose, 0.3% lactose. The optimum fermentation conditions were ：cultivating in 37℃ for 3 h firstly, then culturing in 25℃ for 13 h. Under the optimal condition, the expression level of soluble target protein and bacterial concentration were 2.2 and 2.3 times of those in LB medium under induction of IPTG respectively.