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Soluble Expression and Purification of Bioactive Interleukin 33 in E. coli
Bich Hang Do,박상수,Grace G. Kwon,NGUYEN MINH TAN,강효정,송정아,유지원,Anh Ngoc Nguyen,장재평,장미희,이선주,소성준,심성락,진종화,이경진,Mark J Osborn,최한 한국생물공학회 2017 Biotechnology and Bioprocess Engineering Vol.22 No.3
Interleukin-33 (IL-33) is one of the important alarmins of the immune system and possesses dual functions as an anti- or pro-inflammatory molecule. The production of this cytokine in E. coli is hampered by the insoluble expression in the cytoplasm, resulting in inclusion body formation. In this study, the expression of IL-33 was optimized by fusing the N-terminus of IL-33 with several solubilizing tags that act as chaperones for proper protein folding: maltose binding protein (MBP), b´a´ domain of protein disulfide isomerase (PDIb´a´) and glutathione Stransferase (GST). The expression of the fusion proteins was stimulated by 0.5 mM IPTG at different temperatures, 37, 30, 25, and 18oC. As a result, IL-33 was expressed highly and in soluble form in the cytoplasm of E. coli when fused with MBP or PDIb´a´ tags in the presence of 0.5 mM IPTG at 25 or 30oC. We describe a simple purification procedure of IL-33 from the PDIb´a´-IL-33 construct using immobilized metal affinity chromatography (IMACs) with supplementary of tobacco etch virus (TEV) protease for tag removal. The high bioactivity of purified IL-33 on the proliferation and activation of macrophages was confirmed by MTT and nitrite releasing assays using RAW 264.7 These data show an improved method for producing high grade and yield IL-33.
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