Immunocytochemical and immunochemical study of amelogenesis

采用生化、免疫化学、免疫细胞化学和细胞生物学等方法研究了猪牙胚的成胚性。结果总结如下：1。猪鞘蛋白是一种低分子量蛋白，主要集中在未成熟牙釉质的棱镜鞘中。这些克隆的翻译产物可能是含有395和380个残基的分泌蛋白。我们提出了翻译产物及其蛋白水解裂解产物的术语“鞘层”。全长鞘蛋白与大鼠成釉细胞蛋白氨基酸序列同源性达67%。鞘鞘蛋白的N端裂解肽约有100 ~ 150个氨基酸残基，特异定位于棱镜鞘。c端裂解肽被鉴定为钙结合蛋白，分子量为29kDa和27kda，均可能被磷酸化和糖基化，定位于未成熟釉质的最外层。克隆了编码釉素的cDNA。猪釉素由1104个氨基酸残基组成。分泌的搪瓷素分子量约为150 kDa，定位于靠近成釉细胞质膜的未成熟珐琅质中，与磷灰石晶体无亲和关系。150 kDa的搪瓷蛋白n端裂解蛋白为89 kDa的搪瓷蛋白，对磷灰石晶体具有亲和力。32kDa的搪瓷素氨基酸序列位于89kda搪瓷素的中间部分。综上所述，三类釉质蛋白在成釉发育中起着不同的作用。釉质原蛋白主要形成釉质的形状，釉质素控制釉质初始钙化和磷灰石晶体的生长，鞘蛋白形成棱柱鞘，可能是釉质钙化过程中釉质蛋白逃逸的途径。

Amelogenesis of the porcine tooth germ was investigated by means of biochemical, immunochemical, immunocytochemical and cell biological methods. The results are summarized as follows.1.The cDNAs encoding porcine sheath proteins, which are low molecular weight proteins concentrated in the prism sheath in immature enamel, were cloned and characterized. The translation products of these clones might be secreted proteins having 395 and 380 residues. We proposed the term "sheathlin" for the translation products and their proteolytic cleavage products. Full length sheathlin and rat ameloblastin shear 67% identity in their amino acid sequences.2.The N terminal cleaved peptides of sheathlin had about 100 to 150 amino acid residues and were specifically localized in the prism sheath. The C-terminal cleaved peptides were identified as calcium binding proteins having molecular weights of 29kDa and 27 kDa, both of which might be phosphorylated and glycosylated and were localized outer most layr of immature enamel.3.The cDNA encoding enamelin was cloned. The porcine enamelin composed 1104 amino acid residues. Secreted enamelin having molecular weight of about 150 kDa were localized in immature enamel just adjacent to the plasma membrane of the ameloblast, and showed no affinity for apatite crystals. The N-terminal cleaved protein of the 150 kDa enamelin was 89 kDa enamelin which showed affinity for apatite crystals. The amino acid sequence of the 32kDa enamelin was located in the middle portion of the 89 kDa enamelin.4.In conclusion, three categories of enamel proteins play different roles in the amelogenesis. Amelogenin primarily form the shape of enamel, enamelin control the initial calcification and the growth of the apatite crystals, and the sheathlin produce prism sheaths which might act as a way through which enamel protein escape during enamel calcification.