Prokaryotic expression, purification and activity analysis of recombinant human interleukinª²29

    LI Mingª²Cai, HE Shaoª²Heng

    Allergy and Inflammation Research Institute, Medical College, Shantou University, Shantou 515041, China

    ¡¾Abstract¡¿ AIM: To express recombinant human interleukinª²29 (ILª²29) with biological activity in E. coli. METHODS: The cDNA fragment coding for mature ILª²29 protein was amplified by PCR and cloned into vector pETª²44 Ek/LIC to construct fusion expression vector pETª²44 Ek/LICª²ILª²29. After pETª²44 Ek/LICª²ILª²29 was transformed into E.coli BL21(DE3), the bacteria were induced by IPTG. The expressed ILª²29 fusion protein was purified by Niª²NTA affinity chromatography and the fusion tag was removed from ILª²29 fusion protein by cleavage with enterokinase. The purified ILª²29 was subjected to Nª²terminal sequencing. The antiviral activity of ILª²29 was detected by cytopathic effect reduction assay. RESULTS: The DNA sequencing showed that the expression vector pETª²44 Ek/LICª²ILª²29 was constructed successfully. After induced by IPTG, the target protein accounted for 43% of the total bacterial protein and most was expressed in a soluble form in E. coli cultured at 30¡æ. The purified ILª²29 appeared a single band on SDSª²PAGE and its purity was more than 96%. The first 10 amino acid sequence of the Nª²terminus was consistent with the theoretical sequence. The recombinant ILª²29 showed specific antiviral activity that was comparable to the commercially available IFNª²¦Á2b preparation. CONCLUSION: The recombinant human ILª²29 with biological activity has been obtained. ......