重组大肠杆菌P84A/MC1061诱导和表达卤醇脱卤酶的条件优化

Optimization of inducing and expression conditions for recombinant E. coli P84A/MC1061 to produce halogen alcohol dehalogenases

考察重组工程菌E.coli P84A/MC1061的生长特性,并优化其在卤醇脱卤酶表达过程中的诱导条件和培养条件。在摇瓶发酵中,以阿拉伯糖诱导重组大肠杆菌P84A/MC1061表达卤醇脱卤酶。重点研究培养温度、装液量、摇床转速、培养基初始pH值、诱导时机、诱导剂添加量、诱导剂作用时间对卤醇脱卤酶的表达影响。在培养温度为30℃,装液量为25mL/250mL,摇床转速为200r/min,初始pH值为7.1时,当菌体量OD600值达到2时,按照1‰体积比添加L-阿拉伯糖,诱导培养22h后,发酵液OD值达到21.3,酶活力达到138555.3U/mL。研究发现,培养温度和初始pH值是基因工程菌E.coli P84A/MC1061诱导表达卤醇脱卤酶的显著影响因子,可为卤醇脱卤酶发酵生产的深入研究提供依据。

Growth characteristics of the recombinant genetic engineering bacteria E. coli P84A/MC1061 were investigated and inducing and culttkdng conditions for producing halogen alcohol dehalogenases were optimized. L（＋）- Arabinose was used to induce E. coli P84A/MC1061 to express Halogen alcohol dehalogenases in shaking flasks fermentation. And the study was focused on temperature, filling volume of medium, rotation speed, initial medium pH, inducing timing and adding amount of the inducer was focused in the study. The optimal conditions are as follows： temperature 30℃, medium filling volume 25ml/250ml, rotation speed 200r/min, initial pH value 7.1, and L （＋）- Arabinose was added to fermentation broth when OD600 reached to 2 with the concentration of 1%0. Under the optimal condition, the OD600 and the enzyme activity reached 21.3 and 138555.3U/ml, respectively. The culture temperature and initial medium pH value were significant factors for E. coli P84A/MC1061 to express halogen alcohol dehalogenases.