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Soluble expression and one-step purification of recombinant mouse interferon-lambda 3 in Escherichia coli

机译:重组小鼠干扰素λ3在大肠杆菌中的可溶性表达及一步纯化

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摘要

Interferon (IFN)-lambda 3, a member of the type III IFN family, is a pleiotropic cytokine that exhibits potent antiproliferative, antiviral, and immunoregulatory activities. For further functional study of IFN-lambda 3, we developed an efficient procedure that includes cloning, expression, and purification to obtain relatively large quantity of mouse IFN-lambda 3 fusion protein. The mature IFN-lambda 3 protein-coding region was cloned into the prokaryotic expression vector pET-44. IFN-lambda 3 contains a hexahistidine tag at its C-terminus. We used Ni2+-nitrilotriacetic acid agarose-affinity chromatography to purify the expressed soluble protein. The purified IFN-lambda 3 inhibited significantly IL-13 production in stimulated RAW264.7 macrophages. Our findings show that the production of soluble IFN-lambda 3 proteins by the pET-44 vector in Escherichia coli is a good alternative for the production of native IFN-lambda 3 and could be useful for the production of other IFN proteins.
机译:干扰素(IFN)-λ3是III型IFN家族的成员,是一种多效细胞因子,具有强大的抗增殖,抗病毒和免疫调节活性。为了进一步研究IFN-λ3,我们开发了一种有效的程序,包括克隆,表达和纯化,以获得相对大量的小鼠IFN-λ3融合蛋白。将成熟的IFN-λ3蛋白编码区克隆到原核表达载体pET-44中。 IFN-λ3在其C端含有一个六组氨酸标签。我们使用Ni2 +-亚硝基三乙酸琼脂糖亲和色谱法纯化表达的可溶性蛋白。纯化的IFN-λ3在刺激的RAW264.7巨噬细胞中显着抑制IL-13的产生。我们的发现表明,在大肠杆菌中通过pET-44载体生产可溶性IFN-λ3蛋白是生产天然IFN-λ3的良好选择,并且可用于生产其他IFN蛋白。

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