BIOCHEMICAL-CHARACTERIZATION OF THE MOLECULAR INTERACTION BETWEEN RECOMBINANT BASIC FIBROBLAST GROWTH-FACTOR AND A RECOMBINANT SOLUBLE FIBROBLAST GROWTH-FACTOR RECEPTOR

The extracellular domain of human fibroblast growth factor receptor (X C-FGF-R) was expressed in Escherichia coli. The protein was purified t o homogeneity and the interaction with basic fibroblast growth factor (bFGF), its physiological ligand, was examined. Using resins on which bFGF was reversibly bound, we analysed the characteristics of the bind ing between XC-FGF-R and immobilized bFGF. We also investigated the st oichiometry of the binding between XC-FGF-R and recombinant human bFGF (rhbFGF) applying non-denaturing gel electrophoresis, chemical cross- linking followed by SDS/PAGE, and gel-filtration chromatography. In cr oss-linking and gel-filtration chromatography experiments, a 1:1 compl ex between rhbFGF and XC-FGF-R was observed. The complex was separated from the non-complexed proteins using nondenaturing PAGE in the prese nce of 0.1 % Triton X-100. The band corresponding to the complex was r ecognized by specific antibodies directed against bFGF and its recepto r, blotted on poly(vinylidene difluoride) membranes and submitted to s equence and amino acid analysis. The data obtained from these determin ations confirmed the formation of a 1:1 complex between rhbFGF and XC- FGF-R.