Binding Mode Characterization of Osteopontin on Hydroxyapatite by Solution NMR Spectroscopy

Extracellular matrix glycoproteins play a major role in bone mineralization and modulation of osteogenesis. Among these, the intrinsically disordered protein osteopontin (OPN) is associated with the inhibition of formation, growth and proliferation of the bone mineral hydroxyapatite (HAP). Furthermore, post-translational modifications like phosphorylation can alter conformations and interaction properties of intrinsically disordered proteins (IDPs). Therefore, the actual interaction of OPN with a HAP surface on an atomic level and how this interaction is affected by phosphorylation is of great interest. Here, we study the interaction of full-length OPN on the surface of suspended HAP nanoparticles by solution NMR spectroscopy. We report the binding modes of this IDP and provide evidence for the influence of hyperphosphorylation on the binding character and an explanation for the differing roles in biomineralization. Our study moreover presents an easy and suitable option to measure interaction of nanoparticles in a stable suspension with full-length proteins.