Gene expression and activities analysis of a new fusion protein (RGD) 3/tTF.
- Author:
Jiang-Hua YAN
1
;
Gui-Wang YANG
;
Jie-Ping WANG
;
Na WU
;
Guo-Hong ZHUANG
Author Information
1. Cancer Research Center of Medical School, Xiamen University, Xiamen 361005, China. jhyan@xmu.edu.cn
- Publication Type:Journal Article
- MeSH:
Animals;
Blood Coagulation;
drug effects;
Chromatography, Affinity;
Dose-Response Relationship, Drug;
Electrophoresis, Polyacrylamide Gel;
Enzyme-Linked Immunosorbent Assay;
Escherichia coli;
genetics;
Factor Xa;
metabolism;
Gene Expression;
Humans;
Integrin alphaVbeta3;
metabolism;
Mice;
Oligopeptides;
genetics;
metabolism;
Polymerase Chain Reaction;
Protein Binding;
Recombinant Fusion Proteins;
genetics;
metabolism;
pharmacology;
Thromboplastin;
genetics;
metabolism
- From:
Chinese Journal of Biotechnology
2007;23(3):409-412
- CountryChina
- Language:Chinese
-
Abstract:
To develop a new fusion protein (RGD)3/tTF for the therapy of the selective thrombosis of tumor blood vessels. The fused gene (RGD) 3/tTF was reconstructed by PCR, was cloned into vector pET22 b(+), and expressed in E. coli BL21 (DE3). The fusion protein was purified through Nickel-affinity chromatography column. The tTF activity of the fusion protein was detected by clotting assay and F X activation assay. The specific binding of (RGD) 3/tTF to alphavbeta3 was analyzed by indirect ELISA. The recombinant plasmid pET22 b(+)/(RGD)3/tTF was obtained and expressed in E. coli BL21 (DE3). The purified fusion protein could induce blood coagulation, activiate F X. The ability of (RGD) 3/tTF binding specifically to alphavbeta3 was increased by 32%, compared with RGD/tTF. A new fusion protein (RGD) 3/tTF was successfully expressed in E. coli BL21 (DE3). The expressed proteins retained tTF activity and showed a higher binding to alphavbeta3 than that of RGD/tTF.
