Molecular cloning of a human basic fibroblast growth factor receptor cDNA and expression of a biologically active extracellular domain in a baculovirus system

Michael C. Kiefer, Andrew Baird, Tuan Nguyen, Carlos George-Nascimento, Owen B. Mason, Leslie J. Boley, Pablo Valenzuela, Philip J. Barr

Research output: Contribution to journalArticlepeer-review

33 Citations (Scopus)

Abstract

A cDNA clone encoding a human fibroblast growth factor (FGF) receptor was isolated from a hepatoma cell line cDNA library. The cDNA encodes a three immunoglobulinlike-domain FGF receptor that is similar to a human placental FGF receptor cDNA but lacks two amino acids. The variation observed at these two amino acids, also seen in the two immunoglobulinlike-domain FGF-receptors, can be explained by an alternate splicing mechanism. We have used a baculovirus expression system to produce high levels of a soluble, extracellular domain form of the FGF receptor (EC-FGF receptor). Spodoptera frugiperda (Sf9) insect cells infected with recombinant EC-FGF receptor viruses synthesized and secreted an EC-FGF receptor of apparent Mt=58,000. The EC-FGF receptor purified from conditioned media of infected Sf9 cells by lentil lectin affinity chromatography was shown to bind basic FGF with high affinity (Kd= 1-5 nM), to inhibit the binding of radioiodinated basic FGF to its high affinity receptor and to inhibit endothelial cell proliferation. Furthermore, binding of basic FGF to the EC-FGF receptor was shown to be significantly enhanced by heparin. The availability of biologically active FGF receptors will allow an analysis of their interaction with members of the FGF family of proteins and viruses of the herpes family that have been shown to use the FGF receptor system for cell entry.

Original languageEnglish
Pages (from-to)115-127
Number of pages13
JournalGrowth Factors
Volume5
Issue number2
DOIs
Publication statusPublished - 1 Jan 1991

Keywords

  • Baculovirus expression
  • Basic fibroblast growth factor receptor
  • Cloning
  • Extracellular domain
  • FGF binding
  • Heparin

ASJC Scopus subject areas

  • Endocrinology
  • Clinical Biochemistry
  • Cell Biology

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