Expression, Purification, and Refolding of Recombinant Fusion Protein hIL-2/mGM-CSF  

作　　者：QIAN WEN LI MA WEI LUO MING-QIAN ZHOU XIAO-NING WANG 

机构地区：[1]Institute of Molecular Immunology, Southern Medical University, Guangzhou 510515, Guangdong, China [2]School of Biosciences and Bioengineering, South China University of Technology, Guangzhou 510641, Guangdong, China

出　　处：《Biomedical and Environmental Sciences》2008年第6期509-513,共5页生物医学与环境科学（英文版）

基　　金：supported by the National Natural Science Foundation of China (30771952);the National Natural Science Foundation of Guangdong Province (07117783);NSFC and the Research Grants Council of Hong Kong Joint Research Scheme (30418003).

摘　　要：Objective To study the activities of interleukin （IL）-2 and granulocyte-macrophage colony-stimulating factor （GM-CSF） （hlL-2/mGM-CSF）. Methods SOE PCR was used to change the linker of the fusion protein for higher activities. The fusion protein was expressed in Escherichia coli （E. coil） BL21 （DE3） in inclusion body （IB） form. After IB was extracted and clarified, it was denatured and purified by affinity chromatography. The protein was refolded by dilution in a L-arginine refolding buffer and refined by anion chromatography. The protein activity was detected by cytokine-dependent cell proliferation assay. Results The expression of hIL-2/mGM-CSF in E. coli yielded approximately 20 mg protein/L culture and the purity was about 90%. The specific activities of IL-2 and GM-CSF were 5.4×10^6 IU/mg and 7.1×10^6 IU/mg, respectively. Conclusion This research provides important information about the anti-tumor activity of hIL-2/mGM-CSF in vivo, thus facilitating future clinical research on hlL-2/mGM-CSF used in immune therapy.Objective To study the activities of interleukin （IL）-2 and granulocyte-macrophage colony-stimulating factor （GM-CSF） （hlL-2/mGM-CSF）. Methods SOE PCR was used to change the linker of the fusion protein for higher activities. The fusion protein was expressed in Escherichia coli （E. coil） BL21 （DE3） in inclusion body （IB） form. After IB was extracted and clarified, it was denatured and purified by affinity chromatography. The protein was refolded by dilution in a L-arginine refolding buffer and refined by anion chromatography. The protein activity was detected by cytokine-dependent cell proliferation assay. Results The expression of hIL-2/mGM-CSF in E. coli yielded approximately 20 mg protein/L culture and the purity was about 90%. The specific activities of IL-2 and GM-CSF were 5.4×10^6 IU/mg and 7.1×10^6 IU/mg, respectively. Conclusion This research provides important information about the anti-tumor activity of hIL-2/mGM-CSF in vivo, thus facilitating future clinical research on hlL-2/mGM-CSF used in immune therapy.

关 键 词：HIL-2/mGM-CSF Fusion protein PURIFICATION REFOLDING 

分 类 号：R392-33[医药卫生—免疫学] R373.2[医药卫生—基础医学]