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Growth Factors. 1997;14(4):243-56.

Characterization of a soluble vascular endothelial growth factor receptor-immunoglobulin chimera.

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  • 1Oncology Section, Wyeth-Ayerst Research, Pearl River, New York 10965, USA.

Abstract

To investigate the interaction between vascular endothelial growth factor (VEGF) and its receptor, we have constructed a chimeric protein consisting of the extracellular ligand-binding domain of the human VEGF receptor subtype KDR fused to a human IgG1 Fc domain (KDR-Fc). KDR-Fc was expressed in human 293 kidney epithelial cells as a 300-kDa secreted, dimeric glycoprotein that bound 125I-VEGF165 with high affinity (Kd = 150 pM). Unlike the full length cellular receptor, KDR-Fc did not require heparin for 125I-VEGF165 binding, although heparin did stimulate 125I-VEGF165 binding approximately 50 to 100%. Similar results were observed for KDR-Fc expressed in yeast cells. Since yeast do not synthesize heparan sulfate proteoglycans, we conclude that cellular heparan sulfates do not account for the lack of a heparin requirement for 125I-VEGF165 binding to KDR-Fc. The polycationic protein protamine, which inhibits (IC50 = 1 microgram/ml) 125I-VEGF165 binding to bovine aortic endothelial cells and other KDR-expressing cells by blocking heparin interactions, had no effect on the heparin independent component of 125I-VEGF165 binding to KDR-Fc. Protamine does inhibit (IC50 = 1 microgram/ml) the heparin dependent component of 125I-VEGF165 binding to KDR-Fc. KDR-Fc bound VEGF121 with the same affinity as VEGF165. Heparin had no effect on 125I-VEGF121 binding to KDR-Fc, indicating that heparin interaction with the 44 amino acids contained in VEGF165 but not VEGF121 allow for maximal VEGF165 binding. Deletion analysis of KDR-Fc demonstrated that the determinants required for high affinity VEGF binding are located in the three aminoterminal Ig-domains of the protein. Heparin had no effect on 125I-VEGF165 binding to the three Ig-domain receptor, suggesting that there are heparin binding determinants located in KDR Ig-domains 4 to 7.

PMID:
9386989
[PubMed - indexed for MEDLINE]
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