Immobilization and Clustering of Structurally Defined Oligosaccharides for Sugar Chips:An Improved Method for Surface Plasmon Resonance Analysis of Protein-Carbohydrate Interactions

摘要

Oligosaccharides are increasingly being recognized as important partners in receptor-ligand binding and cellular signaling.Surface plasmon resonance (SPR) is a very powerful tool for the real-time study of the specific interactions between biological molecules.We report here an advanced method for the immobilization of oligosaccharides in clustered structures for SPR and their application to the analysis of heparin-protein interactions.Reductive amination reactions and linker molecules were designed and optimized.Using mono-,tri-,or tetravalent linker compounds,we incorporated synthetic structurally defined disaccharide units of heparin and immobilized them as ligands for SPR.Their binding to an important hemostatic protein,von Willebrand factor (vWf),and its known heparin-binding domain was quantitatively analyzed.These multivalent ligand conjugates exhibited reproducible binding behavior,with consistency of the surface conditions of the SPR chip.This novel technique for oligosaccharide immobilization in SPR studies is accurate,specific,and easily applicable to both synthetic and naturally derived oligosaccharides.