Fluorospectrometric analysis of heparin interaction with fibroblast growth factors.
The fluorescence emission of a single tryptophan residue present in both FGF-1 and FGF-2 was used as a structural probe to directly assess the interaction of the growth factors with heparin or beta-cyclodextran tetradecasulfate. About 20-25% of the fluorescence of either FGF-1 or FGF-2 is quenchable, and is dependent on sulfation of the ligands. The quenchable fluorescence is associated with about 20% of total FGF, suggesting the presence of two fluorospectrometric forms of the protein. The equilibrium dissociation constants, determined by this method, for heparin or beta-cyclodextrin tetradecasulfate binding to FGF-1 are about 1 nM, whereas the values for FGF-2 are 1 and 23 nM, respectively. The method provides a direct tool to evaluate FGF-ligand interaction and assess the structural integrity of the proteins.
Li
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LY
Department of Protein Chemistry, American Cyanamid Company, Pearl River, NY 10965.
Seddon
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AP
D000818:Animals; D001665:Binding Sites; D003505:Cyclodextrins / Q000737:chemistry / Q000378:metabolism; D016220:Fibroblast Growth Factor 1 / Q000737:chemistry / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000737:chemistry / Q000378:metabolism*; D006493:Heparin / Q000737:chemistry / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D008024:Ligands; D011485:Protein Binding; D013050:Spectrometry, Fluorescence; D013431:Sulfates / Q000737:chemistry; D013552:Swine; D014364:Tryptophan / Q000737:chemistry; D047392:beta-Cyclodextrins
D003505:Cyclodextrins; D008024:Ligands; D013431:Sulfates; D047392:beta-Cyclodextrins; D016222:Fibroblast Growth Factor 2; D016220:Fibroblast Growth Factor 1; C064089:beta-cyclodextrin tetradecasulfate; D014364:Tryptophan; D006493:Heparin