Biomimetic Synthesis of an Enamel-Like Material

描述(申请人提供)：生物技术的最新进展使牙釉质细胞外基质中的蛋白质和蛋白酶变得可用，并有助于在体外研究它们的功能，以及首次尝试在人工环境中合成这种钙化组织。目前这项提议的总体目标是确定在蛋白质引导下开发出一种可与人牙釉质的成分、结构和性能相媲美的材料所需的物理化学条件。这项工作将有助于更好地理解釉原蛋白、剪接变异体LRAP+和两种蛋白酶在离子和分子水平上如何在生物矿化过程中发挥作用，以及如何利用这些信息在体外合成釉质。需要检验的假设是，在特定的物理化学条件下，通过使用全长和拼接的人釉原蛋白与矿化溶液中选定的蛋白酶结合，可以仿生合成釉质类材料的薄层，包括组成、微结构和性能。这一假说将通过以下三个具体目标得到验证：(1)确定促进全长釉原蛋白和剪接变异体LRAP+介导的生物矿化的物理化学条件；(2)确定釉原蛋白诱导的生物矿化是否取决于磷灰石基质的晶体取向；以及(3)在矿化进行的同时，通过使用重组基质金属蛋白酶-20和丝氨酸蛋白酶逐渐降解釉原蛋白基质来合成釉质样材料。

DESCRIPTION (provided by applicant): Recent advances in biotechnology made proteins and proteases of the extra cellular matrix of enamel available and facilitate the study of their function in-vitro as well as allowing the first attempts to synthesize this calcified tissue in an artificial environment. The overall objective of the current proposal is to determine the physical-chemical conditions required for a protein-guided development of a material that is comparable to the composition, structure and properties of human enamel. This work should lead to an improved understanding of how amelogenin proteins, the spliced variant LRAP+ and two proteases function in the biomineralization process on an ionic and molecular level and how this information can be used to synthesize enamel in-vitro. The hypothesis to be tested is that a biomimetic synthesis of thin layers of an enamel-like material, including composition, microstructure and properties is feasible by the use of full-length and spliced human amelogenins, when combined with selected proteases in mineralizing solutions at specific physical-chemical conditions. This hypothesis will be tested by the following three specific aims: (1) To determine the physical-chemical conditions that promote biomineralization mediated by full-length amelogenin and the spliced variant LRAP+; (2) To determine if amelogenin-induced biomineralization depends on the crystallographic orientation of the apatite substrate; and (3) To synthesize an enamel-like material by gradually degrading the amelogenin matrix using recombinant MMP-20 and serine proteinase while mineralization proceeds.