title: [Fusion expression and identification of angiostatin and endostatin in E. coli BL21(DE3)].
abstract: Angiostatin(AS) and endostatin(ES) are both potent endogenous angiogenesis inhibitors, and the combination of AS and ES has been shown to have synergistic antiangiogenic effects. Here we report the fusion protein AS-ES expressed in E. coli which has antiangiogenic effects. At first, AS and ES genes were cloned respectively through RT-PCR, then fusion gene was made through gene splicing ,finally pET-42 (b)/AS-ES expression plasmid was constructed and transduced in E. coli BL21 (DE3). Target protein was in form of inclusion body,the rate of expression was about 14%, and MW about 65KD. Western blotting assay showed expressed protein had specific immune reaction to both the antibodies of AS and ES. The expressed protein which was refolded and purified through heparin affinity chromatography had antiangiogenic effect to vessels on chicken embryo chorioallantoic membrane. The results show that fusion protein AS-ES was expressed successfully in E. coli, and the expressed protein,which was renatured and purified, had immuno-reactivity to anti-AS and anti-ES in Western blotting and angiogenesis inhibition activity.
authors.affiliation: Institute for Viral Disease Control and Prevention, Chinese CDC, Beijing 100052, China.
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Mao
authors.forename: Weibin
authors.initials: W
authors.lastname: Liu
authors.forename: Shihong
authors.initials: S
authors.lastname: Fu
authors.forename: Rende
authors.initials: R
authors.lastname: Li
authors.forename: Guodong
authors.initials: G
authors.lastname: Liang
mesh_terms: D020533:Angiogenesis Inhibitors; D043168:Angiostatins / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002642:Chick Embryo; D043169:Endostatins / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification* / Q000494:pharmacology; D011995:Recombination, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
chemical_list: D020533:Angiogenesis Inhibitors; D043169:Endostatins; D011993:Recombinant Fusion Proteins; D043168:Angiostatins