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Optimization Studies on Prokaryotic Cell Expression of the Human Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL)

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Abstract

The aim of the study was to optimize the in vitro induction and expression of the human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and also study the processes of its denaturation, renaturation, and purification. The pGEX-6P-1/TRAIL114–281 plasmid was induced by isopropyl-β-d-1-thiogalactopyranoside (IPTG) in Escherichia coli BL21 (DE3), and the expressed target protein was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The protein expressed in the form of inclusion body was first denaturalized and then renaturalized by dilution and dialysis technique. GST-rTRAIL114–281 fusion protein was purified by Glutathione-Superflow Resin affinity chromatography and confirmed by Western blot. The molecular weight of GST-rTRAIL expressed in E. coli BL21 (DE3) was approximately 40 kDa. GST-rTRAIL was mainly expressed in the form of inclusion bodies. An optimum expression was induced by IPTG at a concentration of 0.2 mM for 8 h at 37 °C. Glutathione-Superflow Resin affinity chromatography yielded the purified GST-rTRAIL protein which was confirmed by Western blot using anti-GST mouse monoclonal antibody. The optimum prokaryotic cell expression of the human GST-rTRAIL was obtained by 0.2 mM IPTG induction for 8 h at 37 °C. The denatured inclusion body protein can be refolded by dilution and dialysis and purified by Glutathione-Superflow Resin affinity chromatography.

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Acknowledgments

We thank Dr. Ming Zhang, The Liver Transplant Center, Ren Ji Hospital, Shanghai Jiaotong University, for his kind advice. This work was supported by National Natural Science Foundation of China [Grant Number 81272557].

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The authors declare that there are no conflict of interest involved.

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Authors and Affiliations

  1. Department of Urology, Xuzhou Center Hospital, Xuzhou, 221009, Jiangsu, China

    Lin Hao, Zhi-Guo Zhang, Zheng-Duo Shi, Kun Pang, Jun-Jie Zhang, Yang Dong & Cong-Hui Han

  2. Department of Urology, Clinical College, Xuzhou Medical College, Xuzhou, 221009, Jiangsu, China

    Lin Hao, Zhi-Guo Zhang, Zheng-Duo Shi & Cong-Hui Han

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  1. Lin Hao
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  2. Zhi-Guo Zhang
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  3. Zheng-Duo Shi
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  4. Kun Pang
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  5. Jun-Jie Zhang
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  6. Yang Dong
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  7. Cong-Hui Han
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Correspondence to Cong-Hui Han.

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Hao, L., Zhang, ZG., Shi, ZD. et al. Optimization Studies on Prokaryotic Cell Expression of the Human Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL). Cell Biochem Biophys 73, 275–279 (2015). https://doi.org/10.1007/s12013-015-0596-6

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  • DOI: https://doi.org/10.1007/s12013-015-0596-6

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