脲梯度扩张床吸附层析复性重组人γ-干扰素  

Refolding of recombinant human interferon-gamma by expanded bed adsorption chromatography with urea gradient

作　　者：靳挺[1] 关怡新[1] 林东强[1] 姚善泾[1] 

机构地区：[1]浙江大学化学工程与生物工程学系

出　　处：《浙江大学学报（工学版）》2006年第7期1257-1261,共5页Journal of Zhejiang University：Engineering Science

基　　金：国家自然科学基金资助项目(20476093)

摘　　要：针对固定床吸附层析技术用于蛋白质体外重折叠复性时,折叠聚集中间体易聚集造成床层堵塞等问题,将扩张床吸附层析技术应用于重组人γ-干扰素包涵体蛋白质的复性研究,并与脲浓度线性梯度法相结合,考察了脲梯度长度和脲终浓度对重组人γ-干扰素复性的影响.复性后的重组人γ-干扰素的纯度达90%;较长的脲梯度长度和合适的脲终浓度有利于蛋白质的复性;在较优的实验条件下,复性后的重组人γ-干扰素收率为53.7%,比活达9.75×106IU/mg.脲梯度扩张床吸附层析法具有处理量大的优点,能有效地复性重组人γ-干扰素包涵体,并能实现复性和分离包涵体蛋白质过程的集成化.To solve the problems of blocking the bed by aggregates in the refolding of recombinant proteins in vitro using packed bed adsorption chromatography, expanded bed adsorption chromatography （EBA） was introduced as the protein renaturation system. The refolding of recombinant human interferon-γ （rhIFN-γ） inclusion bodies was realized by gradually decreasing the concentration of urea in the EBA column after the denatured rhIFN-γ proteins had bound onto the gel. The effects of urea gradient length and final urea concentration on the refolding were investigated. The purity of the refolded rhIFN-γ was up to 90 %, and high urea gradient length and moderate final urea concentration were favorable for the refolding process. Under optimum conditions, the specific activity of rhIFN-γ was up to 9.75 × 10^6 IU/mg and protein recovery was up to 53.7 %. The results showed that the denatured rhIFN-γ was refolded efficiently by EBA with urea gradient, which had the unique capacity for processing large amounts of samples with the refolding and separation process being carried out simultaneously.

关 键 词：重组人Γ-干扰素 包涵体 复性 脲梯度 扩张床吸附层析 

分 类 号：Q511[生物学—生物化学]