DIFFERENCES IN THE INTERACTION OF HEPARIN WITH ARGININE AND LYSINE AND THE IMPORTANCE OF THESE BASIC-AMINO-ACIDS IN THE BINDING OF HEPARIN TO ACIDIC FIBROBLAST GROWTH-FACTOR

Although the interaction of proteins with glycosaminoglycans (GAGs) su ch as heparin are of great importance, the general structural requirem ents for protein- or peptide-GAG interaction have not been well charac terized. Electrostatic interactions between sulfate and carboxylate gr oups on the GAG and basic residues in the protein or peptide dominate the interaction, but the thermodynamics of these electrostatic interac tions have not been studied, Arginine residues occur frequently in the known heparin binding sites of proteins. Arginine is also more common than lysine in randomly synthesized 7-mer peptides that bind to immob ilized heparin and heparan sulfate, We have used heparin affinity chro matography, equilibrium dialysis, and isothermal titration calorimetry techniques to further investigate these interactions. A 7-mer of argi nine eluted from a heparin-affinity column at 0.82 M NaCl, whereas the analogous 7-mer of lysine eluted at 0.64 M, Similarly, the putative h eparin binding site peptide (amino acid residues 110-130) from acidic fibroblast growth factor, which contained four lysine and two arginine residues, eluted at 0.50 M, whereas the analogous peptide with six ly sine residues eluted at 0.41 M and one with six arginine residues elut ed at 0.54 M. At 25 degrees C in 10 mM sodium phosphate, pH 7.4, carbo xy and amino termini blocked arginine (blocked arginine) bound to hepa rin twice as tightly as blocked lysine as measured by equilibrium dial ysis, Similarly, at 30 degrees C in 10 mM sodium phosphate, pH 7.4, an d in water, blocked arginine bound 2.5 times more tightly to anions in heparin than blocked lysine. Using titration calorimetry, the enthalp y of blocked arginine and lysine binding to heparin was 1.14 +/- 0.24 and 0.45 +/- 0.35 kJ/mol, respectively, under identical conditions. Ou r observations show that blocked arginine- and arginine-containing pep tides bound more tightly to GAGs than the analogous lysine species and suggest that the difference was due to the intrinsic properties of th e arginine and lysine side chains, The greater affinity of the guanidi no cation for sulfate in GAGs is probably due to stronger hydrogen bon ding and a more exothermic electrostatic interaction. This can be rati onalized by soft acid, soft base concepts. (C) 1995 Academic Press, In c.