β PDGFR-IgG chimera demonstrates that human β PDGFR Ig-like domains 1 to 3 are sufficient for high affinity PDGF BB binding

To localize human β PDGFR binding determinants, we constructed a fusion protein comprising β PDGFR Ig-like domains 1 to 3 and an IgG₁ Fc domain PDGFR-HFc)- β PDGFR-HFc was expressed as a 200 kDa dimeric molecule and contained Fc epitopes as demonstrated by anti-mouse Fc antibody recognition. Scatchard analysis revealed that PDGF BB possessed a dissociation constant of 1.5 nM for β PDGFR-HFc. Thus, β PDGFR Ig-like domains 1 to 3 are sufficient for high affinity PDGF BB binding. We exploited this fusion protein technology to identify and characterize β PDGFR antagonists using a sensitive β PDGFR immunosorbent assay. In this assay, β PDGFR-HFc half- maximally bound to PDGF BB with an affinity of around 150 pM. Suramin, as well as bacterially expressed and refolded human α PDGFR domains 1-3, inhibited β PDGFR-HFc binding to PDGF BB half-maximally at 25 μM and 10 nM respectively. Therefore, a PDGFR D1-3, like β PDGFR D1-3, are sufficient for high affinity PDGF BB binding. Furthermore, the β PDGFR-HFc immunosorbent assay will be useful to identify β PDGFR antagonists as well as to study α and β PDGFR substitution mutants which further map receptor binding determinants.