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

M. A. Heidaran, D. Mahadevan, W. J. Larochelle

Research output: Contribution to journalArticlepeer-review

21 Scopus citations

Abstract

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.

Original languageEnglish (US)
Pages (from-to)140-145
Number of pages6
JournalFASEB Journal
Volume9
Issue number1
StatePublished - Jan 1 1995
Externally publishedYes

Keywords

  • fusion proteins
  • platelet-derived growth factor receptor
  • suramin

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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