标题：Soluble Expression and Purification of the Functional Interleukin-30 Protein in Escherichia coli

作者：Zhang J1, Liu X, Huang N, Hu Z, Wu W, Teng X, Wang Z, Wei X, Tang H, Wu X, Chen Z, Li J, Li Z.

网址:PMID: 26176652

求助者：lovevv

• 求助时间：2015/7/31 8:52:33
• 求助状态：AB图书馆客服已找到全文，详情咨询在线客服qq 1257749646
• 文献摘要：

Interleukin-30 (IL-30), or IL-27p28, is the α subunit of IL-27 constructed by Epstein-Barr virus induced gene 3 (EBI3) and IL-27p28 binding via non-covalent bonds. IL-30 can be independently secreted and function independent of IL-27. Recent studies demonstrated IL-30 could concurrently antagonize T helper 1 (Th1) and Th17 responses and might have therapeutic implications for controlling autoimmune diseases. However no reports have stated an efficient method to generate relatively large quantity of IL-30. In the present study, anE.coli expression system for the rapid expression of the mouse IL-30 is developed. For the first time, IL-30 was expressed in a form of soluble fusion protein and purified using a way of simple affinity chromatography. In order to avoid the impact of minor codons on expressing eukaryotic protein in E.coli and improve the expression quantity, the nucleotide sequence of IL-30 was optimized. The optimized gene sequence was then sub-cloned into the pET-44a (+) vector, which allowed expression of IL-30 with a fusion tag - NusA. The vector was transformed into E.coli and the expressed fusion protein, NusA-IL-30, was purified by Ni-chromatography. Then the fusion tag was removed by cleavage with thrombin. The purify of purified IL-30 was identified using SDS-PAGE as well as HPLC and the purify was up to about 92%. And the yield of IL-30 was 8.95 mg from 1 L of bacterial culture. Western blot confirmed the identity of the purified protein. The recombinant IL-30 showed its biological activity by inhibiting Th17 differentiating from Naïve CD4+ T cells. Therefore, this method of express and purify IL-30 provide novel procedures to facilitate structural and functions studies of IL-30.

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