Acidic amino acid-rich sequences as binding sites of osteonectin to hydroxyapatite crystals.

Osteonectin, an acidic noncollagenous protein of bone and dentin, has affinity to hydroxyapatite crystals. Binding sites to hydroxyapatite of this protein were determined by a proteolytic experiment and an in vitro binding experiment using synthetic peptide analogues. Osteonectin was adsorbed on hydroxyapatite crystals and digested with trypsin. A peptide was left adsorbed on the crystal even after the digestion. The peptide was identified as an amino terminal peptide containing glutamic acid-rich sequences, which have been assumed to be possible hydroxyapatite-binding sites. Poly glutamic acid sequences were synthesized as models of the binding sites. Glu6 peptide was bound to the hydroxyapatite with a dissociation constant of 2.4 microM. Peptides containing fewer glutamic acids had lower affinity to the crystal. Effects of these peptides on in vitro mineralization were examined by a gel system in microtiter plates. The Glu6 peptide had a positive effect on the mineralization in this system, whereas Asp6 peptide had a negative effect. These effects indicate the presence of an interaction between these peptides and mineral crystals.