Expression of Tumstatin_(183-230)-TRAIL fusion protein and identification of its biological functions
- VernacularTitle:Tumstatin_(183-230)-TRAIL融合蛋白的克隆表达及其生物学双功能鉴定
- Author:
Na REN
;
Liang-Hua WANG
;
Yun GAO
;
Ming-Juan SUN
;
Yu-Liang JIAO
;
Ai-Yun GUO
;
Bing-Hua JIAO
;
- Publication Type:Journal Article
- Keywords:
Tumstatin_(183-230);
TRAIL;
fusion protein;
biology activity
- From:
Academic Journal of Second Military Medical University
1985;0(05):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To express Tumstatin_(183-230)-TRAIL fusion protein and to observe its biological functions.Methods: SOE-ing PCR was employed to amplify the recombinant sequence of Tumstatin_(183-230)and TNF-related apoptosis-inducing ligand (TRAIL_(114-281)).An expression vector pMAL-Tu-T was constructed by inserting Tu-T sequence into pMAL-c_2;the vector was used to transfect E.coli BL21(DE3)and expression of MBP-Tu-T fusion protein was induced by IPTG.Amylose Resin columns were employed to purify the fusion protein.The biological functions of MBP-Tu-T protein was examined by inhibitory test of endothelial cell proliferation,standard tumor cell cytotoxic assay,in vitro tube formation inhibition,and electron microscopic observation(apoptosis).Results:The expression rate of MBP-Tu-T fusion protein in E.coli was about 20%. Purified recombinant protein obviously inhibited endothelial cell proliferation(IC_(50)12.5?g/ml),induced apoptosis of pancreatic cancer cells,and inhibited tube formation.Conclusion:Constructed MBP-Tu-T fusion protein is bifunctional,which lays a solid foundation for further investigation of antitumor effect of Tumstatin_(183-230)-TRAIL in vivo.
