Metal Binding to Sodium Heparin Monitored by Quadrupolar NMR

Heparins and heparan sulfate polysaccharides are negatively charged glycosaminoglycans and play important roles in cell-to-matrix and cell-to-cell signaling processes. Metal ion binding to heparins alters the conformation of heparins and influences their function. Various experimental techniques have been used to investigate metal ion-heparin interactions, frequently with inconsistent results. Exploiting the quadrupolar (23)Na nucleus, we herein develop a (23)Na NMR-based competition assay and monitor the binding of divalent Ca(2+) and Mg(2+) and trivalent Al(3+) metal ions to sodium heparin and the consequent release of sodium ions from heparin. The (23)Na spin relaxation rates and translational diffusion coefficients are utilized to quantify the metal ion-induced release of sodium ions from heparin. In the case of the Al(3+) ion, the complementary approach of (27)Al quadrupolar NMR is employed as a direct probe of ion binding to heparin. Our NMR results demonstrate at least two metal ion-binding sites with different affinities on heparin, potentially undergoing dynamic exchange. For the site with lower metal ion binding affinity, the order of Ca(2+) > Mg(2+) > Al(3+) is obtained, in which even the weakly binding Al(3+) ion is capable of displacing sodium ions from heparin. Overall, the multinuclear quadrupolar NMR approach employed here can monitor and quantify metal ion binding to heparin and capture different modes of metal ion-heparin binding.