Identification of the adsorbing site of lysozyme onto the hydroxyapatite surface using hydrogen exchange and 1H NMR.

The lysozyme-hydroxyapatite interaction was studied by measuring individual hydrogen-deuterium (H-D) exchange rates of amide protons. The H-D exchange reaction was initiated by transferring the lysozyme adsorbed on hydroxyapatite powder from H2O into D2O. After various H-D exchange time periods (pH 7.0, 25 degrees C), the complex was dissociated and the remaining hydrogen label was determined by 2D NMR analysis. The H-D exchange rate of amide protons of residues 9, 11, 13, and 83 was slowed in the hydroxyapatite-lysozyme complex compared with free lysozyme. Residues 9, 11 and 13 positioned at the back of the active site would be the location of the binding site.