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: 7060029
• 负责人: Wendy J Shaw
• 金额: $ 40.49万
• 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/7060029
• 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.

描述：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-功能关系。牙釉质是最高度有序的生物矿化晶体，具有处理磨损和机械应力的独特设计。与许多生物矿化过程一样，尽管蛋白质的存在被认为是至关重要的，但人们对控制牙釉质成核和生长的机制知之甚少。釉质、簇绒蛋白、成釉细胞蛋白和成釉原蛋白都存在于釉质形成过程中，它们都被认为是晶体成核的候选物质。在牙釉质生长过程中，90%的蛋白质都是由淀粉原蛋白组成的，它是牙釉质形成所必需的，也可能是磷酸钙晶体形成的主要因素。除了可能的成核作用外，淀粉原蛋白形成独特的自组装纳米球，这被认为与釉质晶体在发育过程中的拉长生长有关。结构-功能关系将主要通过固态核磁共振（SSNMR）、石英晶体微平衡（QCM）和恒定组成动力学（CCK）来阐明，以研究在发育中的牙釉质中发现的各种条件下固定蛋白。SSNMR将用于准确测定蛋白质或蛋白质自组装与磷酸钙结合的二级结构、动力学和重要的氨基酸。QCM将用于研究成核速率和蛋白质结合动力学。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。位点定向突变，显示导致缺陷的牙釉质也将进行研究，以进一步帮助了解发展中的牙釉质界面。将SSNMR结果与动力学测量相结合，将对釉质基质形成中淀粉原蛋白二级和四级结构的重要性提供大量的见解。这些研究将深入了解牙齿形成过程中的分子水平过程，更广泛地说，将为蛋白质/晶体相互作用提供基本的见解。