title: Binding of bone sialoprotein, osteopontin and synthetic polypeptides to hydroxyapatite.
abstract: The phosphorylated acidic glycoproteins bone sialoprotein (BSP) and osteopontin (OPN) bind to hydroxyapatite (HA) crystals and may be involved in the regulation of bone mineralization. The HA-binding properties of these proteins have been attributed to glutamic acid-rich sequences in BSP and aspartic acid-rich sequences in OPN. The present study examines the roles of these polycarboxylate sequences in the binding of BSP and OPN to HA. Porcine BSP, OPN and the synthetic polypeptides poly-L-glutamic acid [Poly(Glu)] and poly-L-aspartic acid [Poly(Asp)] were labeled with fluorescein isothiocyanate and their binding to HA determined by fluorimetry. From the binding isotherms, dissociation constants (KDs) for all the reagents tested were determined to be in the micromolar range. The saturation binding capacities of HA for Poly(Glu), Poly(Asp), BSP and OPN were similar (500-600 micrograms/m2). To investigate the role of glutamic acid-rich and aspartic acid-rich sequences in the binding to HA of BSP and OPN, respectively, competitive binding studies with Poly(Glu) and Poly(Asp) were performed. Poly(Glu) was able to displace a maximum of 100% of Poly(Glu), 81% of OPN, 68% of BSP and 65% of Poly(Asp). Poly(Asp) was able to displace a maximum of 100% of Poly(Glu), 99% of Poly(Asp), 95% of OPN and 89% of BSP. These results are consistent with the view that BSP and OPN bind to HA via their polycarboxylate sequences, but suggest a complex mode of interaction between polyelectrolytes and ionic crystals.
authors.affiliation: Division of Oral Biology and Department of Biochemistry, School of Dentistry, University of Western Ontario, London, Ontario, Canada N6A 5C1. hagoldbe@julian.uwo.ca
authors.forename: H A
authors.initials: HA
authors.lastname: Goldberg
authors.forename: K J
authors.initials: KJ
authors.lastname: Warner
authors.forename: M C
authors.initials: MC
authors.lastname: Li
authors.forename: G K
authors.initials: GK
authors.lastname: Hunter
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D017886:Durapatite / Q000378:metabolism*; D058574:Integrin-Binding Sialoprotein; D053495:Osteopontin; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism*; D011099:Polyglutamic Acid / Q000138:chemical synthesis / Q000378:metabolism*; D011485:Protein Binding; D012795:Sialoglycoproteins / Q000378:metabolism*; D013552:Swine
chemical_list: D058574:Integrin-Binding Sialoprotein; D010455:Peptides; D012795:Sialoglycoproteins; D053495:Osteopontin; D011099:Polyglutamic Acid; C017645:polyaspartate; D017886:Durapatite