PROTEOLYTIC ACTIVITY OF OPOSSUM TOOTH EXTRACTS

Amelogenins are the main component of the developing enamel matrix. In placental mammals, amelogenins are rapidly cleaved following their se cretion, HPLC fractionation of tooth extracts produces a complex chrom atographic profile. The fractions are rich in amelogenin cleavage prod ucts that generally retain the amino-terminus of the parent protein bu t have varying lengths of peptide removed from the original carboxyl-t erminus. In contrast, HPLC fractionation of opossum tooth extracts pro duces a simple profile with a single major chromatographic peak. SDS-a nd Western blot analyses demonstrated that most of the amelogenin cons isted of a prominent protein band that migrated at 28 kDa. Mass spectr oscopy confirmed the presence of two uncleaved, alternatively spliced forms of opossum amelogenin, Op202 and Op57, but did not detect major amelogenin cleavage products evident in toc,th extracts from placental mammals. Amino acid composition analysis supported the conclusion tha t uncleaved amelogenin is the major component in the developing enamel matrix. Enzymogram analyses using gelatin, casein and recombinant ame logenin as substrates, comparing porcine, rat and opossum tooth extrac ts? suggested that fewer proteinases are present in opossum. These res ults identify potentially significant differences in the proteolytic p rocessing of amelogenins between metatherian and eutherian mammals.