Prokaryotic expression, purification and activity analysis of recombinant human interleukin-29

AIM: To express recombinant human interleukin-29 (IL-29) with biological activity in E. coli. METHODS: The cDNA fragment coding for mature IL-29 protein was amplified by PCR and cloned into vector pET-44 Ek/LIC to construct fusion expression vector pET-44 Ek/LIC-IL-29. After pET-44 Ek/LIC-IL-29 was transformed into E.coli BL21(DE3), the bacteria were induced by IPTG. The expressed IL-29 fusion protein was purified by Ni-NTA affinity chromatography and the fusion tag was removed from IL-29 fusion protein by cleavage with enterokinase. The purified IL-29 was subjected to N-terminal sequencing. The antiviral activity of IL-29 was detected by cytopathic effect reduction assay. RESULTS: The DNA sequencing showed that the expression vector pET-44 Ek/LIC-IL-29 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℃. The purified IL-29 appeared a single band on SDS-PAGE and its purity was more than 96%. The first 10 amino acid sequence of the N-terminus was consistent with the theoretical sequence. The recombinant IL-29 showed specific antiviral activity that was comparable to the commercially available IFN-α2b preparation. CONCLUSION: The recombinant human IL-29 with biological activity has been obtained.