Predimerization of recombinant platelet-derived growth factor receptor extracellular domains increases antagonistic potency

Platelet-derived growth factor (PDGF) is a dimeric growth factor acting thr ough tyrosine kinase alpha- and beta-receptors. In both receptors, the extr acellular parts are composed of five Ig-like domains. Functional mapping of the extracellular part of the receptors have shown that ligand-binding occ urs to Ig-like domains 2 and 3 and that Ig-like domain 4 is involved in rec eptor-receptor interactions. Recombinant GST-fusion proteins of PDGF alpha- receptor Ig-like domains 1-4 and beta-receptor Ig-like domains 1-3 (alpha R D1-4-GST and beta RD1-3-GST) were generated and compared with their cleaved counterparts (alpha RD1-4 and beta RD1-3) with regard to their ability to block PDGF binding to cell surface receptors. In the case of both the alpha - and the beta-receptors, 100-1000-fold lower concentrations of the GST-fus ion proteins were required, as compared to the cleaved forms, for inhibitio n of PDGF binding to cell surface receptors. alpha RD1-4-GST and beta RD1-3 -GST, in contrast to alpha RD1-4 and beta RD1-3, were shown to occur as lig and independent dimers. Covalently cross-linked alpha RD1-4 dimers displaye d a 50-fold increased potency as compared to alpha RD1-4. We thus conclude that the dimeric nature of alpha RD1-4-GST and beta RD1-3-GST is responsibl e for the high antagonistic potency of the fusion proteins.