Purification of EGF-TCS recombinant fusion protein and its targeting action on human tumor cells in vitro
- VernacularTitle:融合蛋白EGF-TCS的纯化及体外靶向杀伤肿瘤细胞的研究
- Author:
Yongmei LI
;
Jinlong CHEN
;
Haiwen YANG
;
Ren LUO
- Publication Type:Journal Article
- Keywords:
epidermal growth factor;
Trichosanthin;
purification;
target action;
apoptosis
- From:Journal of Third Military Medical University
2003;0(13):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To express and purify EGF-TCS fusion protein and observe the targeted and selective killing effect on cancer cells of the fusion protein.Methods The recombinant expression plasmid PQE30/EGF-TCS was transformed to E.coli.M15 and the fusion protein(EGF-TCS) was expressed.Ni-NTA Agrose affinity chromatography was used to purify the protein,flow cytometry to detect EGFR expression rate in cancer cells(BEL-7402,MCF-7,BGC-823) and normal liver LO2 cells,and the killing test to verify selective killing ability of EGF-TCS;The cell apoptosis detection by flow cytometry and microscopic observation were used to confirm the selective killing ability of EGF-TCS.Results Recombinant expression plasmid PQE30/EGF-TCS was expressed in E.coli.M15 stably and effectively.The purity of EGF-TCS was over 95% by chromatography.EGFR expression rate was highest in hepatoma cells BEL-7402(72.33%) and lowest in normal liver LO2 cells(5.51%).The killing ability of recombinant protein was more effective to cancer cells(IC50 of BEL-7402,MCF-7 and BGC-823 was 11.4,22.47 and 12.53 ?g/ml respectively) and was weak to normal cells(IC50 53.19 ?g/ml).Conclusion The recombinant protein EGF-TCS that induces apoptosis of cancer cells was successfully constructed by gene engineering technology.
