Expression and characterization of fusion protein RGD/tTF for targeting therapy of cancer / 中国免疫学杂志

ABSTRACT

Objective:To express and purify a new fusion protein(RGD/tTF) for targeting therapy of cancer, and analyze its activities.Methods:The fused gene RGD/tTF was constructed by PCR, and then was inserted into vector pET22 b(+),expressed in E.coli BL21(DE3).The fusion protein was purified through Nickel-affinity chromatography column. The tTF activity of the fusion protein was analyzed by clotting assay and FⅩ activation assay. The specific binding of RGD/tTF to ?_v?_3 was analyzed by indirect ELISA.Results:The recombinant plasmid pET22 b(+)/RGD/tTF with correct sequence was obtained. The fusion protein was expressed with high yield in E.coli BL21(DE3). The purified fusion protein could activiate FⅩ and cause blood coagulation, and bind to ?_v?_3 specifically.Conclusion:The recombinant plasmid pET22 b(+)/RGD/tTF was constructed.The fusion protein retained TF activity and binding specificity to ?_v?_3, lays the foundation for studying the function of inducing thrombosis in tumor vasculature in vivo.