Improving the mda-7/IL-24 refolding and purification process using optimized culture conditions based on the structure characteristics of inclusion bodies  

作　　者：Xiaojuan Wang Chaogang Bai Jian Zhang Aiyou Sun Xuedong Wang Dongzhi Wei 

机构地区：[1]State Key Laboratory of Bioreactor Engineering,East China University of Science and Technology,130 Meilong Road,Shanghai 200237,People’s Republic of China

出　　处：《Bioresources and Bioprocessing》2014年第1期82-90,共9页生物资源与生物加工(英文)

基　　金：supported by a grant from the Ministry of Science and Technology(National Major Science and Technology Projects of China(No.2012ZX09304009)).

摘　　要：Background:The melanoma differentiation-associated gene-7(mda-7)/interleukin-24(IL-24)can induce apoptosis in a wide variety of tumor cell types,whereas it has no toxicity in normal cells.However,recombinant human mda-7/IL-24 is difficult to obtain from Escherichia coli because of its insolubility.Results:In this study,we improved the structure of inclusion bodies(IBs)by optimizing the induction temperature,pH,concentrations of inducer,and metal ion additives.Statistically designed experimental analyses of three metal ion factors were performed using the Box-Behnken design.Induction temperature of 30℃,pH 7.0,and 0.1 mM isopropyl-β-D-thiogalactopyranoside(IPTG)were selected,and the optimized levels for the factors predicted by the model comprised the following:Mg^(2+)(15.7 mM),Ca^(2+)(16.6 mM),and Mn^(2+)(3.0 mM).The optimized culture conditions improved the structure of the IBs,which was validated by scanning electron microscopy(SEM)and the increase of IBs solubility.Conclusions:After optimization,IB solubility and renatured mda-7/IL-24 increased by 51%and 84%,respectively.This study also provided a simple purification method of specific IB washing steps.Manipulating the fermentation parameters to optimize the refolding and purification process is likely to be widely applicable to other proteins.

关 键 词：Escherichia coli EXPRESSION Inclusion body INTERLEUKIN-24 PURIFICATION REFOLDING 

分 类 号：R73[医药卫生—肿瘤]