重组耐碱性蛋白A大肠杆菌工程菌株的发酵优化-《集美大学学报（自然科学版）》

文章信息/Info

Title:: Fermentation Optimization of Recombinant Alkaline Resistant Protein A Engineering Strain of Escherichia coli

作者:: 黄玲珑1; 蓝萌萌1; 倪辉2; 李志朋1; 3; 洪菲菲4; 洪清林4; 姜泽东1; 3; 朱艳冰1; 3; 1.集美大学海洋食品与生物工程学院，福建厦门 361021；2.厦门海洋职业技术学院，福建厦门 361102；3.福建省食品微生物与酶工程重点实验室，福建厦门 361021；4.绿新（福建）食品有限公司，福建漳州 363199

Author(s):: HUANG Linglong1; LAN Mengmeng1; NI Hui2; LI Zhipeng1; 3; HONG Feifei4; HONG Qinglin4; JIANG Zedong1; 3; ZHU Yanbing1; 3; 1.College of Ocean Food and Biological Engineering,Jimei University,Xiamen 361021,China;2.Xiamen Ocean Vocational College,Xiamen 361102,China;3.Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering,Xiamen 361021,China;4.Greenfresh (Fujian) Foodstuff Co.,Ltd.,Zhangzhou 363199,China

Keywords:: recombinant alkali-resistant protein A; Escherichia coli; optimization of fermentation media; optimization of fermentation conditions

摘要:: 对重组耐碱性蛋白A的大肠杆菌工程菌株进行发酵优化以提高目标蛋白表达量，筛选出TB培养基作为基础发酵培养基；通过单因素实验在摇瓶水平上对培养基组分进行优化，然后在最优培养基的基础上对发酵条件进行优化，最后通过30 L发酵罐进行放大验证。结果表明，最优培养基组分为蔗糖8 g/L、大豆蛋白胨12 g/L、酵母浸粉24 g/L、 KH2PO423.1 g/L、K2HPO4 125.4 g/L ，pH =7.0；最优摇瓶发酵条件为5%接种量、40%装液量、37 ℃培养至工程菌株A(600)=0.8时加入0.3 mmol/L IPTG、37 ℃诱导8 h，优化后的蛋白A表达量为优化前的3.01倍。30 L发酵罐放大验证结果表明，37 ℃诱导8 h后，蛋白A表达量达到最高，与摇瓶水平优化结果一致。

Abstract:: Fermentation conditions were optimized for the engineered Escherichia coli strain expressing of recombinant alkaline resistant protein A in order to enhance the expression level of the target protein.TB medium was identified as the most suitable basial fermentation medium through screening.The composition of the culture medium was optimized at the shaker flask scale through single-factor experiments.Subsequently,fermentation conditions were optimized using the improved medium.Finally,scale-up validation was performed using a 30 L fermenter.The results showed that the optimal medium components were sucrose 8 g/L,soybean peptone 12 g/L,yeast extract 24 g/L,KH2PO4 23.1 g/L,K2HPO4 125.4 g/L,and pH =7.0.The optimal shaker flask fermentation conditions were 5% inoculation volume,40% liquid loading volume,incubation at 37 ℃ until the the engineered strain A(600)=0.8,then adding 0.3 mmol/L IPTG,and induction at 37 ℃ for 8 h.The expression level of protein A was 3.01 times than that before optimization.The results of 30 L fermenter amplification indicated that the expression of protein A reached the highest level after induction at 37 ℃ for 8 h,which was consistent with the optimization results at the shaker flask level.