Solid State NMR Structure/Function Studies of Amelogenin

釉原蛋白的固态核磁共振结构/功能研究

• 批准号: 6926515
• 负责人: Wendy J Shaw
• 金额: $ 37.07万
• 依托单位: BATTELLE PACIFIC NORTHWEST LABORATORIES
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6926515
• 关键词: amelogenin; calcium phosphate; chemical kinetics; crystallization; dental development; molecular assembly /self assembly; nuclear magnetic resonance spectroscopy; protein binding; protein structure function; site directed mutagenesis; structural biology

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.

产品说明：本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与功能关系。牙釉质是最高度有序的生物矿化晶体，并且被独特地设计为处理磨损和机械应力。与许多生物矿化过程一样，尽管蛋白质的存在被认为是至关重要的，但人们对控制釉质成核和生长的机制知之甚少。釉质蛋白、tuftelins、成釉蛋白和釉原蛋白都存在于釉质形成过程中，并且都被认为是晶体成核的候选物。釉原蛋白由釉质生长过程中存在的90%的蛋白质组成，是适当的釉质形成所必需的，并且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之外，釉原蛋白形成独特的自组装纳米球，其被认为与发育期间釉质晶体的伸长生长有关。结构-功能关系将主要使用固态NMR（SSNMR），石英晶体微天平（QCM）和恒定组成动力学（CCK）来阐明，以研究在发育釉质中发现的各种条件下的固定化蛋白质。SSNMR将用于准确确定二级结构，动力学和氨基酸结合蛋白质或蛋白质自组装磷酸钙的重要性。QCM将用于研究成核速率和蛋白结合动力学。将用CCK探测表面特异性相互作用，以确定晶体生长抑制和晶体生长机制的变化。还将研究导致牙釉质缺陷的定点突变，以进一步帮助了解发育中的牙釉质界面。将SSNMR结果与动力学测量相关联，将为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提供大量的见解。这些研究将深入了解牙齿形成中涉及的分子水平过程，更一般地说，将提供对蛋白质/晶体相互作用的基本见解。