Characterization of vascular endothelial cell growth factor interactions with the kinase insert domain-containing receptor tyrosine kinase - A real time kinetic study

The kinase insert domain-containing receptor (KDR) tyrosine kinase mediates calcium mobilization in endothelial cells and plays a key role during phys iological and pathological angiogenesis, To provide a detailed understandin g of how KDR is activated, we analyzed the kinetics of ligand-receptor inte raction using BIAcore. Both predimerized (KDR-Fc) and monomeric (KDR-cbu) r eceptors were examined with vascular endothelial cell growth factor (VEGF) homodimers and VEGF/placental growth factor (PIGF) heterodimers. VEGF binds to KDR-Fc with k(a) = 3.6 +/- 0.07e(6), k(d) = 1.54 +/- 0.19e(-4), and K-D = 37.1 +/- 4.9 pM. These values are similar to those displayed by monomeri c KDR where k(a) = 5.23 +/- 1.4e(6), k(d) = 2.74 +/- 0.76e(-4), and KD = 51 .7 +/- 5.8 pM were apparent. In contrast, VEGF/PIGF bound to KDR-Fc with k( a) = 7.3 +/- 1.6e(4), k(d) = 4.4 +/- 1.2e(-4), and K-D = 6 +/- 1.2 nM. Thus , the heterodimer displays a 160-fold reduced K-D for binding to predimeriz ed KDR, which is mainly a consequence of a 50-fold reduction in k(a). We we re unable to detect association between VEGF/PIGF and monomeric KDR. Howeve r, nanomolar concentrations of VEGF/ PIGF were able to elicit weak calcium mobilization in endothelial cells. This latter observation may indicate par tial predimerization of KDR on the cell surface or facilitation of binding due to accessory receptors.