title: Partial characterization of endothelial FGF receptor functional domain by monoclonal antibody VBS-1.
abstract: Polypeptide growth factors mediate their cellular responses by binding to and activating specific cell surface receptors. Monoclonal antibody (MAb) VBS-1, produced against native fibroblast growth factor receptor-1 (FGFR-1), inhibited the binding of fibroblast growth factor-2 (FGF-2) to its receptor on coronary venular endothelial cells (CVECs) as determined by 125I-FGF-2 Scatchard analysis and [3H]thymidine uptake assays (ED50 = 80 ng/mL). Enzyme studies demonstrated that MAb VBS-1 binds to a protein epitope. Proteolytic mapping of the CVEC-FGFR established that a 52 kDa doublet contained the FGF binding site and the MAb VBS-1 antigenic epitope. N-glycanase digestion suggested the presence of a 50 kDa core protein for the CVEC-FGFR. Tunicamycin treatment resulted in the loss of expression of the core protein and the mature receptor, indicating the importance of CVEC-FGFR n-linked glycosylation. By Northern blot analysis, it was determined that CVECs express fgfr-1 and not fgfr-2. VBS-1 recognized FGFR-1 (140 kDa) and crossreacted weakly with FGFR-2 (135 kDa). Using a combination of affinity crosslinking, proteolytic mapping and Mab VBS-1 binding studies, we have located the FGF binding site near the NH2-terminal domain of the receptor close to the highly acidic box.
authors.affiliation: Department of Genetics and Cell Biology, Washington State University, Pullman, WA 99164, USA. vblanck@univ-lemans.fr
authors.forename: Vincent D
authors.initials: VD
authors.lastname: Blanckaert
authors.forename: Subramaniam
authors.initials: S
authors.lastname: Venkateswaran
authors.forename: In Suk
authors.initials: IS
authors.lastname: Han
authors.forename: Kwan Hee
authors.initials: KH
authors.lastname: Kim
authors.forename: Michael D
authors.initials: MD
authors.lastname: Griswold
authors.forename: Margaret E
authors.initials: ME
authors.lastname: Schelling
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D000939:Epitopes / Q000276:immunology; D006825:Hybridomas; D011233:Precipitin Tests; D017434:Protein Structure, Tertiary; D017468:Receptors, Fibroblast Growth Factor / Q000032:analysis* / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D017468:Receptors, Fibroblast Growth Factor