CHARACTERIZATION DMP1 A DENTIN PHOSPHOPROTEIN

骨骼和牙齿的适当矿化对于 正常的人体生长发育和肌肉骨骼功能。的 非胶原基质蛋白的作用和机制 （NCP）的牙质，特别是那些专门起源于 对成牙本质细胞的了解还不够。最重要的角色可能是 在牙本质形成和修复性牙本质形成期间表达， 牙本质，并可能包括矿化过程的调节。在 尽管多年的研究，非常复杂的主要牙本质NCP 没有被详细描述。然而，我们最近完成了 这些NCP之一的cDNA测序其染色体定位， 原位杂交，获得的证据表明，其发育调控 在表达上。这种蛋白质，首先通过克隆命名来识别， AG 1是第一个被测序和定位的牙本质特异性蛋白 染色体上。这项研究具有重要的临床应用价值。DMP 1映射 在人类染色体上与牙本质形成基因相同的区域 II型（DI-II）。在DI-II中，牙本质不完全矿化 是最重要的病理发现基本假设是， 非胶原蛋白如Dmp 1与胶原基质相互作用， 特定位点，成核并调节矿化过程。我们现在 建议从大鼠基因组文库中鉴定Dmp 1基因，并确定 基因5 '端的序列，从而鉴定启动子 这对于理解Dmp 1的组织特异性表达是必要的。 为了了解蛋白质的功能，我们建议 大量的重组蛋白。这方面的工作有两个目标： l）制备未修饰的脱辅基蛋白，使得其可以用作 用于研究翻译后修饰的底物， 磷酸化和糖基化; 2）制备多克隆抗体。这 抗-Dmp 1抗体将用于检查Dmp 1在 修复性和套质以及Dmp 1在 细胞外基质在发育过程中。免疫电镜 程序和金结合的DMP 1抗体将用于 定位，并确定分泌DMP 1的途径，从 成牙本质细胞这些实验将提供更多的了解 Dmp 1合成的分子机制 成牙本质细胞长期目标是了解Dmp 1在 牙本质形成

The proper mineralization of bones and teeth has great importance in normal human growth and development and musculo-skeletal function. The roles and mechanisms of action of the non-collagenous matrix proteins (NCPs) of dentin and particularly those originating specifically in the odontoblasts are not well understood. The most important roles may be expressed during the periods of dentinogenesis and formation of reparative dentin, and may include regulation of the process of mineralization. In spite of many years of study the very complex principal dentin NCPs have not been characterized in detail. However we have recently completed the cDNA sequencing of one of these NCPs its chromosomal localization, and, by in situ hybridization, gained evidence that its developmentally regulated in expression. The protein, first identified by the cloning designation, AG1, is the first dentin-specific protein to be sequenced and localized chromosomally. This study has important clinical applications. Dmp1 maps to the same region on the human chromosome as the gene for dentenogenesis imperfecta type II (DI-II). In DI-II incomplete mineralization of dentin is the most important pathological finding. The basic hypothesis is that noncollagenous proteins like Dmp1 interacts with the collagen matrix at specific loci, nucleate and regulate the process of mineralization. We now propose to identify the Dmp1 gene from a rat genomic library and determine the sequences at the 5'end of the gene and thus identify the promoter which is necessary to understand the tissue specific expression of Dmp1. In order to understand the function of the protein we propose to make large amounts of recombinant protein. Work on this aim has two objectives: l) to prepare the unmodified apoprotein so that it can be used as a substrate for studying post-translational modifications like phosphorylation and glycosylation, 2) to prepare polyclonal antibody. This anti-Dmp1 antibody would be used to examine the presence of Dmp1 in reparative and mantle dentin as well as localization of Dmp1 in the extracellular matrix during development. Immunoelectron microscope procedures and gold-conjugated DMP1 antibody would be used for localization and to determine the pathway for secretion of DMP1 from the odontoblasts. These experiments will provide increased understanding of the molecular mechanisms involved in the synthesis of Dmp1 by the odontoblasts. The long -term goal is to understand the role of Dmp1 in dentinogenesis.