Prokaryotic Expression,Purification and Bioactivity of Recombinant Mouse Interleukin 17F/His Fusion Protein

Objective To express mouse interleukin 17F(mIL-17)/His fusion protein in E.coli,purified the expressed product and analyze its bioactivity.Methods The mIL-17f gene was amplified by RT-PCR and inserted into prokaryotic expression vector pET28a.The constructed recombinant plasmid pET28a /mIL-17f was transformed to E.coli BL21(DE3)was transformed to E.coli BL21(DE3)for expression under induction of IPTG.The expressed rmIL-17F /His fusion protein was purified by Ni2+-NTA affinity chromatography and identified for reactogenicity.BALB/c mice were immunized with the purified and re-naturalized rmIL-17 /His fusion protein by nasal drip,then determined for expression of IL-6 mRNA in lung tissue by real-time fluorescent quantitative PCR,and for IL-17F,IL-4 and IFNγ levels in peripheral blood by ELISA.Results The DNA sequence of amplified mIL-17f gene was consistent with that reported in GenBank.Recombinant plasmid pET28a /mIL-17f was constructed correctly.The expressed recombinant protein,with a relative molecular mass of about 19 000,mainly existed in a form of inclusion body and contained about 30% of total somatic protein.The purified fusion protein reached a purity of about 95% and showed good reactogenicity.The mucosal immunization with the purified fusion protein promoted the expression of IL-6 mRNA in lung tissues and increased the IL-4 and IFNγ levels in sera of mice.Conclusion The rmIL-17F with bioactivity was highly expressed in E.coli and purified,which laid a foundation of further study on the function of IL-17F.