Characterization studies of the acquired enamel pellicle formed in vitro

Abstract

The purpose of this study is the identification and characterization of those salivary proteins which constitute precursor proteins to the acquired enamel pellicle. The goal of this investigation is to identify proteins in pellicles formed in vitro by the adsorption of pure glandular secretions to hydroxyapatite powder (HA). Either parotid or submandibular secretions were incubated with HA at a ratio of 5 mg/mlof saliva for 24 hours, in the presence of 0.02% NaN3. The suspension was centrifuged at 27,000 x g, and the pellet was washed with 0.2M NaCl, pH 7.5 and stirred at room temperature for 24 hours in 0.2M EDTA pH 7.5 to solubilize the mineral phase. The adsorbed protein was recovered following dialysis against water and lyophilization. The proteins were characterized by means of electrophoresis, using 7.5% polyacrylamide discontinuous gels, gradient SDS-PA gels, and Isoelectric focusing gels. Further characterization was carried out by means of C18 reversed phase high performance liquid chromatography. The amino acid compositions of these proteins were also determined. These studies led to the identification of specific phosphoproteins, namely proline-rich proteins, statherin, and histatins as major constituents of the in vitro pellicle derived from both parotid and submandibular saliva. In addition, minor components were identified in submandibular derived pellicle.