Expression of biologically active isoforms of the tumor angiogenesis factor VEGF in Escherichia coli.
Vascular endothelial growth factor (VEGF) was identified as an endothelial cell specific mitogen that induces angiogenesis and vascular permeability in vivo. VEGF is a homodimeric protein which contains three intramolecular and two intermolecular disulfide bridges. Here, we report on an efficient procedure for recombinant production of VEGF isoforms VEGF121 and VEGF165 in Escherichia coli. The proteins were solubilized from inclusion bodies, refolded, and purified by chromatographic methods. The final protein products were almost completely in the dimeric conformation, bound to VEGF receptor FLT1 with a Kd of 30 pM, stimulated proliferation of human umbilical vein endothelial cells half-maximally at a concentration of 30 pM, and induced in vivo neovascularization and vascular permeability on the chicken chorioallantoic membrane.
Siemeister
G
G
Tumor Biology Center, Institute of Molecular Medicine, Freiburg Federal Republic of Germany.
Schnurr
B
B
Mohrs
K
K
Schächtele
C
C
Marmé
D
D
Martiny-Baron
G
G
D000482:Allantois / Q000098:blood supply; D000818:Animals; D001667:Binding, Competitive; D002199:Capillary Permeability / Q000187:drug effects; D002478:Cells, Cultured; D002642:Chick Embryo; D002823:Chorion / Q000098:blood supply; D003001:Cloning, Molecular; D016228:Endothelial Growth Factors / Q000096:biosynthesis* / Q000737:chemistry / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects; D004926:Escherichia coli; D006801:Humans; D007700:Kinetics; D008222:Lymphokines / Q000096:biosynthesis* / Q000737:chemistry / Q000494:pharmacology*; D046911:Macromolecular Substances; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D011487:Protein Conformation; D017510:Protein Folding; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D017978:Receptors, Growth Factor / Q000378:metabolism*; D040262:Receptors, Vascular Endothelial Growth Factor; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000494:pharmacology; D014471:Umbilical Veins; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D008222:Lymphokines; D046911:Macromolecular Substances; D017978:Receptors, Growth Factor; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D020794:Receptor Protein-Tyrosine Kinases; D040262:Receptors, Vascular Endothelial Growth Factor