[The cloning and expression of angiogenic inhibitor tumstatin(45-132) in E. coli].
OBJECTIVE
To clone tumstatin(45-132) gene and to express recombinant human tumstatin(45-132) in E. coli BL21.


METHODS
Tumstatin gene was cloned by RT-PCR and then tumstatin(45-132) gene amplified from tumstatin gene was cloned into pBV220. The recombinant plasmid pBV220-tumstatin(45-132) was sequenced and transformed into E.coli BL21. E. coli BL21 transformed with the recombinant plasmid pBV220-tumstatin(45-132) was induced at 42 degrees Celsius. After the recombinant tumstatin(45-132) was purified, its bioactivity was detected by endothelia cell proliferation test.


RESULTS
264 bp tumstatin(45-132) fragment was cloned and its sequence was identical with that in GenBank. The tumstatin(45-132) was expressed in E. coli BL21. Expressed product accounted for about 10% of total bacterial proteins and its relative molecular mass (M(r)) was 9,600. The purified protein showed inhibitory effect on proliferation of endothelia cells ECV304 in vitro.


CONCLUSIONS
tumstatin(45-132) gene has been cloned and expressed successfully in E.coli BL21. Expressed tumstatin(45-132) can inhibit the proliferation of endothelial cell ECV304.
Luo
Yi-qin
YQ
Department of Biochemistry and Molecular Biology, College of Basic Medical Sciences, Second Military Medical University, Shanghai 200433, China.
Wang
Liang-hua
LH
Qiu
Yi
Y
Jiao
Bing-hua
BH
D020533:Angiogenesis Inhibitors / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D001324:Autoantigens / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D049109:Cell Proliferation / Q000187:drug effects; D003001:Cloning, Molecular; D024141:Collagen Type IV / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D006801:Humans; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology
D020533:Angiogenesis Inhibitors; D001324:Autoantigens; D024141:Collagen Type IV; D011994:Recombinant Proteins; C057112:type IV collagen alpha3 chain