MATRIX PROTEIN CONTROL OF CRYSTAL NUCLEATION AND GROWTH

本项目的目的是确定釉质基质蛋白作为釉质矿物形成潜在调节因子的结构-功能关系。我们的工作假设是，可溶的亲水性釉质蛋白(如釉质蛋白)与不溶的基质成分(如釉原蛋白聚集体)一起提供适当的表面特征来诱导磷酸钙的成核。在适当浓度的矿物离子存在下，这会导致形成非常薄的釉质矿物带。这些釉质带在厚度和宽度上的生长立即被疏水蛋白质(例如溶液中的釉原蛋白)所抑制，这些蛋白质吸收到形成晶体的特定表面上。进一步推测，这些矿物质带在组织成熟过程中的开始生长是由釉质蛋白水解酶降解这些被吸收的蛋白抑制物控制的。为了更好地了解基质蛋白如何控制牙釉质等组织中的矿化，我们更好地了解了基质蛋白如何控制牙釉质等组织中的矿化，我们建议使用重组釉原蛋白和釉质蛋白来测定主要的釉质基质蛋白在羟基磷灰石(HA)和磷酸八钙(OCP)晶体上的吸附等温线。选择HA和OCP作为釉质矿物形成的原型进行研究。我们进一步建议确定吸附的重组釉原蛋白和釉质蛋白对HA种子晶体生长的抑制作用，并将这些发现与蛋白质吸附的程度相关联。我们将进一步评估重组釉质蛋白酶(如釉质蛋白和EMPS1)降解吸附在HA和OCP表面的釉质蛋白的能力，因为这可能是釉质蛋白酶控制矿化的一个重要机制。我们进一步建议使用琼脂糖凝胶和共价连接到琼脂糖珠上的蛋白质来评估重组釉原蛋白和釉蛋白在体外促进磷酸钙成核的能力，并影响晶体的形态。从长远来看，这些研究对开发新的仿生方法可能是重要的，这些方法可以用于治疗受损或患病的牙齿。

The purpose of this project is to determine the structure-function relationship of enamel matrix proteins as potential regulators of enamel mineral formation. Our working hypothesis is that soluble hydrophilic enamel proteins (e.g. enamelin) in association with insoluble matrix components (e.g. amelogenin aggregates) provide the appropriate surface characteristics to induce the nucleation of calcium phosphates. In the presence of appropriate concentrations of mineral ions, this results in the formation of very thin ribbons of enamel mineral. The growth of these enamel ribbons in thickness and width is immediately inhibited by hydrophobic proteins (e.g. amelogenins in solution) which absorb onto specific faces of the forming crystals. It is further hypothesized that the onset of growth of these mineral ribbons during tissue maturation is controlled by the degradation of these absorbed protein inhibitors by enamel proteinases. To gain a better understanding how matrix proteins can control mineralization in a tissue like enamel, we gain a better understanding how matrix proteins can control mineralization in a tissue like enamel, we proposed to determine the adsorption isotherms for the major enamel matrix proteins onto hydroxyapatite (HA) and octacalcium phosphate (OCP) crystals, using recombinant amelogenin and enamelin. HA and OCP were chosen for study as prototypes for forming enamel mineral. We further propose to determine the inhibitory effect of adsorbed recombinant amelogenin and enamelin on HA seeded crystal growth and relate these findings to the extent of protein adsorption. We will further assess the ability of recombinant enamel proteinases (e.g. enameylsin and EMPSl) to degrade enamel proteins which are absorbed onto HA and OCP surfaces, since this may be an important mechanism by which enamel proteinases can control mineralization. We further propose to assess the ability of recombinant amelogenin and enamelin to promote calcium phosphate nucleation in vitro, using agarose gels and proteins covalently linked to sepharose beads, and to influence crystal morphology. Long term, these studies may be important to the development of new biomimetic approaches which could be used to treat damaged or diseased teeth.