摘要
重组基因工程菌经中试发酵后,诱导表达出以包涵体形式存在的重组大肠杆菌不耐热肠毒素B亚单位(LTB)蛋白。采用Q Sepharose High Perfomance阴离子交换层析,将洗涤、裂解后得到的可溶性蛋白进行纯化,得到纯度高达98%的重组LTB蛋白。利用Sephadex G-25分子筛层析法将重组LTB蛋白中的盐和尿素脱去,使蛋白复性,最终获得目的蛋白纯度为95%。将得到的重组LTB蛋白进行免疫双扩散实验,结果表明此蛋白具有良好的生物学活性。通过高效表达重组LTB蛋白的大肠杆菌发酵,确定了其中试发酵工艺,并建立了Q柱一步纯化即可获得目的蛋白的方法,该工艺简捷、高效、易于工业化生产,为重组LTB蛋白的生产和应用奠定了基础。
The recombinant bacteria subunit （LTB） in the form of inclsion body were induced to express Escherichia coli heat labile enterotoxin B after pilot-scale fermentation. Q Sepharose High Perfomance Anion exchange column chromatography were used to purify the soluble protein after processing of washing and lysing, resulting to obtain the recombinant protein LTB with a high purify of 98%. Renaturation the protain through Sephadex G-25 molecular sieve chromatography to remove the salts and urea, and ultimately obtain the target protein of 95% putify. The immune double diffusion test was performed to indicate that the target protein has a good biological activity. This was the first time to determine the optimal pilot-scale fermentation conditions, the production process of E. coli LTB recombinant proteins, and also to establish the method in the use of Q column to obtain the target protein in only one step. The process was more convenient and efficient for industrial production which laid a solid foundation for the production and application of recombinant LTB protein.
出处
《中国生物工程杂志》
CAS
CSCD
北大核心
2015年第2期78-83,共6页
China Biotechnology
基金
国家科技重大专项子课题(2014ZX09102042-002)
安徽省科技攻关计划(1301041020)资助项目
关键词
LTB
包涵体
纯化
LTB Inclusion bodies Purification
