Interaction of soluble and surface-bound heparin binding growth-associatedmolecule with heparin

The interaction of heparin with heparin binding growth-associated molecule (HB-GAM) was studied using isothermal titration calorimetry (ITC) and surfa ce plasmon resonance (SPR), ITC studies showed that, in solution, heparin b ound HB-GAM with a Delta H of -30 kcal/mole corresponding to a dissociation constant (K-d) of 460 nM, The stoichiometry of interaction was 3 moles of HB-GAM per mole of heparin, corresponding to a minimum heparin binding site for HB-GAM of 12-16 saccharide residues. Kinetic measurements of heparin i nteraction with HB-GAM made by SPR afforded a K-d of 4 nM, suggesting consi derably tighter binding when HB-GAM was immobilized on a surface, Affinity chromatography of a sized mixture of heparin oligosaccharides, having a deg ree of polymerization (dp) of >14 saccharide units, on HB-GAM-Sepharose dem onstrated that oligosaccharides having more than 18 saccharide residues sho wed the tightest interaction. (C) 1999 Federation of European Biochemical S ocieties.