Fuantai-03(FAT-03), isolated from the Dasyatis akajei, has a strong antiangiogenic activity. The recombinant Fuantai-03 (GST/rFAT-03) fusion protein can be obtained with the DNA recombination technology. In this study, expression conditions of GST/rFAT-03 were optimized by response surface experimental design method. The constructed engineering bacteria containing GST/rFAT-03 plasmid was induced by isopropy-beta-D-thiogalactosid (IPTG), the GST affinity column was used for isolation and purification, and then the effects of different culture time, IPTG concentration, induction temperature and induction time on the amount of soluble GST/rFAT-03 fusion protein were compared. The culture time for optimal expression was 6.13 h, IPTG concentration was 0.36 mmol/L, induction temperature was 19.71 degrees C, and induction time was 13.60 h. The amount of soluble GST/rFAT-03 fusion protein was 7.57 mg/L under above mentioned expression conditions. The results also showed that rFAT-03 significantly inhibited angiogenesis in chicken chorioallantoic membrane in a dose-dependent manner. Moreover, the soluble form of the target protein is useful for further work on purification and on studying its biological function.
Angiogenesis Inhibitors ; biosynthesis ; genetics ; Animals ; Chickens ; Chorioallantoic Membrane ; blood supply ; Escherichia coli ; genetics ; metabolism ; Fish Proteins ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; Skates (Fish)