NDLI: A novel self-cleavable tag Zbasic–∆I-CM and its application in the soluble expression of recombinant human interleukin-15 in Escherichia coli

Content Provider	Springer Nature Link
Author	Shi, Siwei Chen, Huanhuan Jiang, Hua Xie, Yueqing Zhang, Lei Li, Ninghuan Zhu, Chencen Chen, Junsheng Luo, Han Wang, Jiaxian Feng, Lei Lu, Huili Zhu, Jianwei
Copyright Year	2016
Abstract	Soluble expression of recombinant therapeutic proteins in Escherichia coli (E. coli) has been a challenging task in biopharmaceutical development. In this study, a novel self-cleavable tag Zbasic–intein has been constructed for the soluble expression and purification of a recombinant cytokine, human interleukin-15 (IL-15). We screened several solubilizing tags fused with the self-cleavable Mycobacterium tuberculosis recA mini-intein ∆I-CM and demonstrated that Zbasic tag can significantly improve the solubility of the product with correspondent to the intein activity. The fusion protein “Zbasic–∆I-CM–IL-15” was expressed with high solubility and easily enriched by the cost-effective cation-exchange chromatography. The self-cleavage of the fusion tag Zbasic–∆I-CM was then induced by a pH shift, with an activation energy of 7.48 kcal/mol. The mature IL-15 with natural N-terminus was released and further purified by hydrophobic interaction and anion-exchange chromatography. High-resolution reverse-phase high-performance liquid chromatography and mass spectrometry analysis confirmed that the product was of high purity and correct mass. With a CTLL-2 cell proliferation-based assay, the EC50 was evaluated to be of about 0.126 ng/mL, similar to the product in clinical trials. By avoiding the time-consuming denaturing-refolding steps in previously reported processes, the current method is efficient and cost-effective. The novel tag Zbasic–∆I-CM can be potentially applied to large-scale manufacturing of recombinant human cytokines as well as other mammalian-sourced proteins in E. coli.
Starting Page	1133
Ending Page	1142
Page Count	10
File Format	PDF
ISSN	01757598
Journal	Applied Microbiology and Biotechnology
Volume Number	101
Issue Number	3
e-ISSN	14320614
Language	English
Publisher	Springer Berlin Heidelberg
Publisher Date	2016-09-28
Publisher Place	Berlin, Heidelberg
Access Restriction	Subscribed
Subject Keyword	Intein Zbasic Interleukin-15 Soluble expression C-terminal cleavage Microbiology Microbial Genetics and Genomics Biotechnology
Content Type	Text
Resource Type	Article
Subject	Medicine Applied Microbiology and Biotechnology Biotechnology

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