[Separation and purification of rat immature enamel proteins]

Sprague Dawley rats were chosen to prepare immature and soft enamel tissue. The authors used a sequential dissociative extraction scheme in which the enamel matrix proteins were extracted first in guanidine HCl and then in guanidine HCl-EDTA, just as the method described by Termine, et al. The first step completely extracted amelogenins and the second step got enamelin. These two groups of protein were further purified by Sephadex-G200 gel filtration chromatography and analysed by polyacrylamide gel electrophoresis. The results showed: amelogenins were resolved into three fractions eluted from the column, and expressed as A1, A2, A3, respectively. The main fraction A1, which had a molecular weight range from 18 kD to 29 kD, had a main protein band of 23 kD when estimated by SDS-PAGE. Enamelin was eluted into two fractions called E1 and E2. The main fraction E1 had a single band of 44 kD on the SDS-PAGE gel. PAGE showed that amelogenins had many subunits. The authors also observed that the amelogenins would aggregate and precipitate reversibly when pH value, ionic strength or protein concentration changed.