Sensing ligand binding to a clinically relevant lectin by tryptophan fluorescence anisotropy

Göhler, Antonia; Büchner, Claudia; André, Sabine; Doose, Sören; Kaltner, Herbert und Gabius, Hans-Joachim (2011): Sensing ligand binding to a clinically relevant lectin by tryptophan fluorescence anisotropy. In: Analyst, Bd. 136, Nr. 21: S. 5270-5276

Abstract

Increasing insights into the involvement of endogenous lectins in disease processes fuel the interest to develop potent inhibitors. As a consequence, robust assay procedures are required. Due to their activity as adhesion/growth-regulatory effectors this study focussed on galectins. The human proto-type galectin-1 was selected as representative of this family with conserved presence of a tryptophan moiety in the binding site. This structural feature was taken advantage of to establish its use as reporter for ligand contact measuring polarized fluorescence emission. The experimentally determined anisotropy r0 was about 0.2, altered by about 5% in the presence of the cognate disaccharide lactose. This parameter change enabled calculating the equilibrium binding constant and kinetic rate constants. The detailed analysis of the depolarization process further indicated fast conformational dynamics within the binding site. Since an inherent property of the protein was exploited, no labeling is needed. Owing to tryptophan’s presence in carbohydrate-binding sites, also in other classes of lectins as well as in carbohydrate-binding modules and glycoenzymes (glycosyltransferases, glycosidases), this assay procedure can have relevance beyond galectins.

• Dokumententyp: Zeitschriftenartikel
• Fakultät: Tiermedizin > Veterinärwissenschaftliches Department > Lehrstuhl für Physiologische Chemie
• Themengebiete: 500 Naturwissenschaften und Mathematik > 570 Biowissenschaften; Biologie
• Allianz-/Nationallizenz: Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.
• Sprache: Englisch