Solid State NMR Structure/Function Studies of Amelogenin

描述：这项研究的总体目标是阐明驱动牙釉质形成的生物矿化蛋白的结构-功能关系。牙釉质是最有序的生物矿化晶体，专为处理磨损和机械应力而设计。与许多生物矿化过程一样，尽管蛋白质的存在被认为是关键，但人们对控制牙釉质成核和生长的机制知之甚少。在釉质形成过程中，釉质蛋白、成釉蛋白、成釉蛋白和成釉蛋白都存在，它们都被认为是晶体成核的候选物质。釉原蛋白由牙釉质生长过程中90%的蛋白质组成，是牙釉质正常形成所必需的，也可能是磷酸钙晶体发育的主要贡献者。除了可能的成核作用外，釉原蛋白还形成独特的自组装纳米球，这些纳米球被认为与釉质晶体在发育过程中的拉长生长有关。结构-功能关系将主要使用固体核磁共振(SS核磁共振)、石英晶体微天平(QCM)和恒定组成动力学(CCK)来研究在发育牙釉质中发现的各种条件下的固定化蛋白质。SS核磁共振将被用来准确地确定蛋白质或蛋白质自组装与磷酸钙结合所必需的二级结构、动力学和氨基酸。QCM将用于研究成核速率和蛋白质结合动力学。用CCK研究晶体的表面特异性相互作用，以确定晶体生长的抑制和晶体生长机制的改变。还将研究导致牙釉质缺陷的定点突变，以进一步帮助理解牙釉质界面的发育。将SS核磁共振结果与动力学测量结果相关联，将有助于深入了解釉原蛋白的二级和四级结构在釉质基质形成中的重要性。这些研究将深入了解牙齿形成所涉及的分子水平的过程，更广泛地说，将为蛋白质/晶体相互作用提供基本的见解。

DESCRIPTION: The overall goal of this research is to elucidate the structure-function relationships of the biomineralization proteins driving the formation of enamel. Enamel is the most highly ordered biomineralization crystal and is uniquely designed to handle abrasions and mechanical stress. As with many biomineralization processes, though, very little is understood about the mechanisms controlling enamel nucleation and growth, although the presence of proteins has been deemed critical. Enamelins, tuftelins, ameloblastins and amelogenins are all present during enamel formation and all have been suggested as candidates for crystal nucleation. Amelogenin consists of 90% of the protein present during enamel growth, is necessary for proper enamel formation and is likely a major contributor in the development of the calcium phosphate crystal. In addition to a possible nucleation role, amelogenin forms unique self assembled nanospheres which are thought to be tied to the elongated growth of enamel crystals during development. Structure-function relationships will be elucidated primarily using solid state NMR (SSNMR), Quartz Crystal Microbalance (QCM) and constant composition kinetics (CCK) to study the immobilized protein under the wide variety of conditions found in developing enamel. SSNMR will be used to accurately determine the secondary structure, dynamics and amino acids important in binding the protein or protein self assembly to calcium phosphates. QCM will be used to investigate nucleation rates and protein binding kinetics. The surface specific interactions will be probed with CCK to determine crystal growth inhibition and change in crystal growth mechanism. Site directed mutations, shown to cause defects in enamel will also be studied, to further aid in understanding of the developing enamel interface. Correlating the SSNMR results with kinetic measurements will provide a great deal of insight into the importance of the secondary and quaternary structure of amelogenin in formation of the enamel matrix. These studies will yield an in depth understanding of the molecular level processes involved in the formation of teeth, and more generally will provide basic insight into protein/crystal interactions.