Characterization of the extracellular domain in vascular endothelial growth factor receptor-1 (Flt-1 tyrosine kinase).
Flt-1 tyrosine kinase, vascular endothelial growth factor (VEGF) receptor-1, binds VEGF and a new VEGF-related ligand, placenta growth factor, but KDR/Flk-1 (VEGF receptor-2) binds only VEGF. To characterize the functional regions in the Flt-1 extracellular domain such as the ligand binding region and the dimer formation of the receptor, we constructed a series of mutants of the Flt-1 extracellular domain as soluble forms in a baculovirus system. We found that a region carrying the N-terminal 1st to 3rd immunoglobulin (Ig)-like domains of Flt-1 binds both ligands with high affinity. However, for dimer formation of soluble Flt-1, a region further downstream in the Flt-1 extracellular domain was required. Mutant Flt-1 receptors expressed in COS cells confirmed the requirement of the 4th to 7th Ig region for the activation of Flt-1 tyrosine kinase. Soluble Flt-1 carrying the N-terminal 1st to 3rd Ig region suppressed VEGF-dependent endothelial proliferation in vitro to the same level as the larger forms of soluble Flt-1, suggesting that the binding of one soluble Flt-1 molecule to one subunit of the VEGF homodimer may be sufficient to block the VEGF activity.
Tanaka
K
K
Department of Genetics, University of Tokyo.
Yamaguchi
S
S
Sawano
A
A
Shibuya
M
M
D000818:Animals; D016367:Baculoviridae; D001665:Binding Sites; D019556:COS Cells; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D016228:Endothelial Growth Factors / Q000378:metabolism / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D006801:Humans; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D008222:Lymphokines / Q000378:metabolism / Q000494:pharmacology; D008961:Models, Structural; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D011487:Protein Conformation; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000502:physiology*; D020794:Receptor Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000737:chemistry* / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D017384:Sequence Deletion; D018411:Spodoptera; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A; D040281:Vascular Endothelial Growth Factor Receptor-1; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D008222:Lymphokines; D009838:Oligodeoxyribonucleotides; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D020794:Receptor Protein-Tyrosine Kinases; D040281:Vascular Endothelial Growth Factor Receptor-1