乳糖诱导剂促进P450 BM-3在大肠杆菌中可溶性表达的研究

摘要

P450 BM-3是一种具有工业化应用潜力的单加氧酶，可催化饱和脂肪酸羟基化。为提高其在大肠杆菌宿主中的可溶性表达水平，采用乳糖作为诱导剂对P450 BM-3的诱导表达条件进行研究。结果发现：在大肠杆菌的OD600达到0.7~1.5时，添加2.0 g/L的乳糖、30℃诱导10 h可获得最佳诱导效果。与IP TG的诱导效果对比发现：采用乳糖作诱导剂时，菌体生物量提高1.09倍，目标蛋白量提升2.13倍，蛋白包涵体的比例则降低至10%。研究结果表明：乳糖可显著提升P450 BM-3在大肠杆菌中的重组表达水平，并且能够促进p450 BM-3的可溶性表达。%P450 BM-3 is a monooxygenase with the potential for industrial application. It can catalyze saturated fatty acids hydroxylation. In order to enhance the expression level and the solubility of P450 BM-3 in Escherichia coli BL21 (DE3), studies was carried out with lactose as the inducer. The expressions conditions in shake flask were optimized. It was found that while 2. 0 g/L lactose was added into the culture (OD600=0. 7~1. 5) and incubated at 30℃ for 10 hours, the yield of P450 BM-3 arrived at the maximum (80 mg/L). With lactose induction, the biomass was doubled, the amount of P450 BM-3 achieved 626. 8 nmol/L, which was 3. 13 times of that with IPTG induction. The solubility of P450 BM3 was also improved, for the inclusion bodies decreased to 10%. These data indicated that lactose was effective to improve the expression of P450 BM-3 in E. coli and would be conducive to the soluble expression of P450 BM-3.