人白细胞介素4的原核表达、复性、纯化及生物学活性鉴定  

Prokaryotic Expression, Purification,Refolding and Biological Assays of Recombinant Human Interleukin 4

作　　者：李炯[1] 崔凯军[1] 文静[1] 赵志伟[1] 陈平[1] 田聆[1] 阚兵[1] 文艳君[1] 邓洪新[1] 范琳玉[1] 魏于全[1] 

机构地区：[1]四川大学华西医院生物治疗国家重点实验室

出　　处：《生物医学工程学杂志》2007年第4期866-869,共4页Journal of Biomedical Engineering

基　　金：国家自然科学基金资助项目(30300313)

摘　　要：用PCR方法从pORF-hIL4质粒中扩增重组人白细胞介素4(hIL-4)的基因,构建PQE60原核表达载体并诱导其目的蛋白的表达。对表达的目的蛋白的可溶性进行鉴定,发现hIL-4基因表达产物在大肠杆菌中主要以不溶性包涵体形式存在,经过超声波破细菌、包涵体提取、洗涤处理后,用5mol/L盐酸胍变性溶解包涵体沉淀,上清稀释复性后透析,再经镍亲和层析进行纯化。纯化后的hIL-4进行TF-1细胞体外增生实验检测其生物学活性后,可用于人外周血树突状细胞的体外诱导培养。A DNA fragment encoding human interleukin 4 was obtained by PCR from pORF-hIL4 plasmid. The amplified fragment was inserted into prokaryotic expression vector PQE60 and recombinant protein was expressed in E. Coli M15 by adding isopropyl-β-D-thiogalactoside （IPTG）. The hIL-4 protein was present as insoluble inclusion bodies in the bacterial extract. After denaturation of inclusion bodies with 5 mol/L guanidine hydrochloride, the supernate was diluted to get renaturized. Then dialysis and Ni chelating chromatography were used for purification. TF-1 proliferation assay of recombinant human interleukin 4 was performed ,and then rhIL-4 was fit to be used for proliferation of human dendritic cells from monocyte in vitro.

关 键 词：重组人白细胞介素4 包涵体 复性 纯化 生物学活性 树突状细胞 

分 类 号：R392[医药卫生—免疫学]