重组人白细胞介素22的量化生产及生物学活性鉴定

Objective: To optimize the conditions for heterologous production of human interleukin-22 (IL-22) in E. coli and to functionally test its biological activity in cultured human hepatic cell line and in a mouse model of liver disease. Methods: An expression plasmid containing the human full-length IL-22 cDNA fused in-frame with the Small Ubiquitin-like Modifier (SUMO) tag was introduced into the E.coli strain BL21(DE3) through heat shock transformation. The transformed BL21 (DE3) cells were cultured at 18, 22 and 37℃and IL-22 expression was induced using different concentrations of IPTG. Re-natured and soluble IL-22 was purified by affinity chromatography and functionally tested in cultured human hepatic cell line HepG2 and in a mouse model of spontaneous liver inflammation. Results: At all three induction temperatures, the obtained recombinant IL-22 protein, with a molecular mass of approximately 16.7 kDa, was mainly present in the form of inclusion body (IB). Following a 4-step denaturating/renaturing protocol and affinity chromatography, 130mg/L of soluble IL-22 protein with purity over 90% was obtained. When added to cultured HepG2 cells, recombinant IL-22 at a concentration of 100-4000 ng/mL showed detectable mitogenic activity. And when injected intraperitoneally into mice with spontaneous hepatitis, IL-22 was found to exacerbate liver inflammation. Conclusion: Functionally active human IL-22 fusion protein can be efficiently produced in E. coli. The therapeutic value of recombinant IL-22 and economic viability of the production system developed was discussed in detail in the current study.