PHOSPHOPROTEIN ANALYSIS OF SEQUENTIAL EXTRACTS OF HUMAN DENTIN AND THE DETERMINATION OF THE SUBSEQUENT REMINERALIZATION POTENTIAL OF THESE DENTIN MATRICES

Phosphoprotein appears to play an important role in the mineralization of dentin during tooth development and remineralization after deminer alization by dental caries. To better understand this role, we describ e the extraction and characterization of phosphoprotein from immature, human root apex dentin during and after EDTA demineralization. The ex traction procedure included dissociation of the demineralized dentin m atrix by guanidine hydrochloride (Gdn . HCl) followed by subsequent di gestion with cyanogen bromide (CNBr) and collagenase. Characterization of these extracts included 'Stains-All' staining of SDS polyacrylamid e gels (SDS-PAGE) and amino acid, protein and phosphorus analyses. The ability of these matrices to remineralize was determined by TEM and m easuring calcium levels in the remineralized tissue by atomic absorpti on spectroscopy. The staining of SDS-PAGE gels and amino acid analysis showed that an intact phosphophoryn was extracted from the dentin of the immature apices during EDTA demineralization and that it had an ap parent M-r similar to 140,000. In the subsequent extracts and digests, the phosphoprotein has a range of molecular weights, some of which ma y have been degraded products of the intact phosphoprotein. A greater quantity of phosphoprotein was found in the EDTA-demineralized dentin matrices than in dentin after Gdn . HCI, CNBr and collagenase digests. These EDTA-demineralized matrices also remineralized to a greater ext ent than those dissociated with Gdn . HCI. The differences in both the quantity and the quality, as defined by the amino acid residue profil e, of the phosphoprotein in the sequential extracts of the root apex d entin may be important in affecting the ability of this tissue to remi neralize.