Enamel, the hardest and most mineralized tissue in the human body, is comprised of a unique organization of apatite nanofibers of only 50 nm width but several micrometers to millimeters in length. Its structure is the result of a protein-guided uniaxial growth process of apatite crystals along their c-axes in a three-dimensional organic framework that hydrolyzes in coordination with advancing mineralization to transform into a tissue almost entirely comprised of mineral. While the role of self-asembly of enamel matrix proteins, in particular amelogenin, has widely been recognized as a crucial factor in controlling structure development of enamel, the current model based on the formation of amelogenin nanospheres has significant limitations with regards to the ability of a spherical structure guidin the anisotropic growth of initially ribbon-like apatite crystals and their transformation into a compact mineralized structure. Amelogenin is a hydrophobic protein which comprises about 90% of the enamel matrix proteins. Recently we discovered that the recombinant human full-length amelogenin protein (rH174) forms ribbons of 17 nm width, which grow over a period of days to several micrometer in length. Such ribbons have the ability to self-align and to form bundles which resemble the appearance of aligned apatite crystallites in an enamel rod. Ribbon formation requires the presence of both calcium and phosphate ions suggesting that ion bridges develop and drive the self-assembly process. Synthesis of such ribbons was kinetically enhanced in a water-oil emuslion system but ribbons were also generated in an oil-free environment and commonly formed within 3 to 5 days of incubation in calcium phosphate solutions. While nanoribbons of rH174 contain calcium and phosphate, they do not directly promote apatite crystllization, but instead appear to stabilize an amorphous mineral. Oriented apatite formed however on nanoribbons made from an amelogenin cleavage product, rH146, indicating that the processing of the full- length protein might induce a transformation from amorphous to crystalline apatite. This proposal is based on the hypothesis that amelogenin nanoribbons are the biologically relevant supramolecular structures in developing enamel and hydrolysis of nanoribbons is required to enable oriented calcium phosphate mineralization. This hypothesis will be tested through the following two specific aims: 1. To induce oriented calcium phosphate crystal growth on amelogenin nanoribbons in-vitro;2. To demonstrate that amelogenin nanoribbons, as observed in-vitro, are the predominant supramolecular structure of developing enamel in-vivo.
The current model of how amelogenin proteins organize mineralization in dental enamel is inconclusive. This study is based on the observation that amelogenin the prominent constituent of the enamel matrix can form nanoribbons in-vitro. The studies are designed to provide evidence of such ribbons in-vivo in developing enamel of mice and will lead to a change in the current paradigm of protein controlled enamel mineralization.
|Carneiro, Karina M M; Zhai, Halei; Zhu, Li et al. (2016) Amyloid-like ribbons of amelogenins in enamel mineralization. Sci Rep 6:23105|
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