Angiogenic activity of a fusion protein of the cell-binding domain of fibronectin and basic fibroblast growth factor.
We constructed a fusion protein of the cell-binding domain of human fibronectin and human basic fibroblast growth factor, and prepared a polypeptide with both cell-adhesive activity and growth factor activity. A human gene fragment coding for basic fibroblast growth factor was amplified by the polymerase chain reaction, and introduced into the expression vector pTF7520, which encodes the cell-binding domain of human fibronectin. The resulting plasmid encoded a fusion protein in which basic fibroblast growth factor was added covalently to the C-terminal end of the fibronectin fragment. The fusion protein was expressed in Escherichia coli JM109 cells and purified from the extract by heparin affinity chromatography. The purified fusion protein had cell-adhesive activity toward BALB/c 3T3 cells, and stimulated their DNA synthesis in serum-depleted cultures. The fusion protein gave maximum mitogenic activity at the concentration of 10 nM. The fusion protein adsorbed to culture dishes, or added to collagen gels, stimulated the growth of human umbilical-vein endothelial cells. The fusion protein stimulated the angiogenesis in chorioallantoic membranes of developing chick embryos.
Hashi
H
H
Biology Division, National Cancer Center Research Institute, Tokyo, Japan.
Hatai
M
M
Kimizuka
F
F
Kato
I
I
Yaoi
Y
Y
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002448:Cell Adhesion / Q000187:drug effects; D002642:Chick Embryo; D004247:DNA / Q000096:biosynthesis; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects; D016222:Fibroblast Growth Factor 2 / Q000737:chemistry*; D005353:Fibronectins / Q000737:chemistry*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009389:Neovascularization, Pathologic / Q000139:chemically induced*; D010957:Plasmids; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D014471:Umbilical Veins / Q000166:cytology / Q000187:drug effects
D005353:Fibronectins; D011993:Recombinant Fusion Proteins; D016222:Fibroblast Growth Factor 2; D004247:DNA