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

Platelet-derived growth factor (PDGF) is a dimeric growth factor acting through tyrosine kinase alpha- and beta-receptors. In both receptors, the extracellular parts are composed of five Ig-like domains. Functional mapping of the extracellular part of the receptors have shown that ligand-binding occurs to Ig-like domains 2 and 3 and that Ig-like domain 4 is involved in receptor-receptor interactions. Recombinant GST-fusion proteins of PDGF alpha-receptor Ig-like domains 1-4 and beta-receptor Ig-like domains 1-3 (alphaRD1-4-GST and betaRD1-3-GST) were generated and compared with their cleaved counterparts (alphaRD1-4 and betaRD1-3) 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-fusion proteins were required, as compared to the cleaved forms, for inhibition of PDGF binding to cell surface receptors. alphaRD1-4-GST and betaRD1-3-GST, in contrast to alphaRD1-4 and betaRD1-3, were shown to occur as ligand independent dimers. Covalently cross-linked alphaRD1-4 dimers displayed a 50-fold increased potency as compared to alphaRD1-4. We thus conclude that the dimeric nature of alphaRD1-4-GST and betaRD1-3-GST is responsible for the high antagonistic potency of the fusion proteins.