Effect of Otoconial Proteins Fetuin A, Osteopontin, and Otoconin 90 on the Nucleation and Growth of Calcite

Abstract

We investigated the roles of three proteins associated with the formation of otoconia including fetuin A, osteopontin (OPN), and otoconin 90 (OC90). In situ atomic force microscopy (AFM) studies of the effects of these proteins on the growth of atomic steps on calcite surfaces were performed to obtain insight into their effects on the growth kinetics. We also used scanning electron microscopy to examine the effects of these proteins on crystal morphology. All three proteins were found to be potent inhibitors of calcite growth, although fetuin A promoted growth at concentrations below about 40 nM and only became an inhibitor at higher concentrations. We then used in situ optical microscopy to observe calcite nucleation on films of these proteins adsorbed onto mica surfaces. By measuring the calcite nucleation rate as a function of supersaturation, the value of the interfacial energy that controls the free energy barrier to heterogeneous nucleation was determined for each protein. OPN and OC90 films led to significantly reduced interfacial energies as compared to the value for homogeneous calcite nucleation in bulk solution. The value for fetuin A was equal to that for bulk solution within experimental error. Zeta potential measurements showed all of the proteins possessed negative surface charge and varied in magnitude according to sequence fetuin A > OC90 > OPN. In addition, the interfacial energies exhibited an inverse scaling with the zeta potential. In analogy to previous measurements on polysaccharide films, this scaling indicates the differences between the proteins arise from the effect of protein surface charge on the solution–substrate interfacial energy.

The role of phosphorylation and dephosphorylation of shell matrix proteins in shell formation: anin vivoandin vitrostudy

“…It is assumed that the inhibition of calcite steps is due to step-specific binding affinity that results in shape modification, similar to the effect of otoconial proteins on the calcite crystallization. 39 Strikingly, in the presence of 10 μg mL −1 dephosphorylated proteins, the shape modification effects on crystals were largely neutralized ( Fig. 5B1, B2, D1 and D2).…”

“…Step patterns on the faces of the crystals indicated rounding of one certain calcite step and hindering the calcium or carbonate ions from attaching to the kink sites, which is usually associated with preferential binding of proteins to those steps. 39 Then, after dephosphorylation, the shape modification ability of the SMPs was compromised, as is evident from the rhombohedral calcium carbonate formed in the presence of dephosphorylated EISMs (Fig. 6B1, B2, D1 and D2).…”

Functional derivatives of human dentin matrix protein 1 modulate morphology of calcium carbonate crystals

“…Interestingly, a lot of proteins involved in the biomineralization of calcium phosphate and calcium carbonate are IDPs . These include, for example, mammalian proteins: osteopontin (OPN), bone sialoprotein (BSP), dentin sialophosphoprotein (DSPP), as well as a zebrafish otolith matrix protein Starmaker (Stm), and its homologs from other fish species . However, more ordered proteins are also involved, for example: otoconin‐90 (OC90), amorphous calcium carbonate‐binding protein (ACCBP), otolin‐1 …”

Otoconia and otolithic membrane fragments within the posterior semicircular canal in benign paroxysmal positional vertigo

“…Otoconin‐90 accounts for 90% of the total protein in the organic core of the otoconia, and together with otolin, forms iso‐oriented columns of nano‐crystals, which then results in the complex mosaic of native otoconia . Fetuin‐A and osteopontin are also believed to regulate nucleation and growth of calcite . Otopetrin 1 has been demonstrated to regulate cytosolic concentrations of calcium during otoconial mineralization.…”

“…16,19 Fetuin-A and osteopontin are also believed to regulate nucleation and growth of calcite. 20 Otopetrin 1 has been demonstrated to regulate cytosolic concentrations of calcium during otoconial mineralization. Otoconia have been shown to exhibit degenerative changes with age.…”