Expression of recombinant N-glycosylation interleukin 24 and study of inducing tumor cells apoptosis in vitro / 中国药理学通报

ABSTRACT

Aim To obtain recombinant human IL-24 secretorily expressed in Pichia pastoris,and study the activity of inducing tumor cells apoptosis of this N-glycosylation protein. Methods By the recombinant plasmid ?/pUC18,the confirmed IL-24 gene was inserted between the sites of BamH Ⅰ and EcoR Ⅰ of expression plasmid pPIC9K. The recombinant plasmid IL-24/pPIC9K was transformed into P. pastoris strain GS115. Yeast transformants were induced for expression of recombinant human IL-24 with methanol. The desired protein was identified with Tricine-SDS-PAGE and Western blot.Amount of IL-24 was assayed with ELISA and the glycosylation was analyzed by PNGase F.The activity of inducing tumor cells apoptosis was confirmed by MTT assay and Hoechst assay in vitro.Results Recombinant expression plasmid IL-24/pPIC9K was successfully constructed. 5 transformants were screened with G418 and PCR. Induced with methanol,the expression level of IL-24 was (81.31?14.46) mg?L-1 at flask fermentation,and 70 % IL-24 generated N-glycosylation.Recombinant IL-24 induced apoptosis in MCF-7 cells,but not in NHLF.Conclusion The secretorily expression of the N-glycosylation IL-24 protein in P. pastoris and the study of inducing tumor cells apoptosis lay the foundation for the further study of molecular mechanism of IL-24 on anti-tumors and the potential application.