A chondroitin sulfate chain attached to the bone dentin matrix protein 1 NH2-terminal fragment
A chondroitin sulfate chain attached to the bone dentin matrix protein 1 NH2-terminal fragment
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DOI：
10.1074/jbc.m512964200
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发表时间：
2006-03-24
影响因子：
4.8
通讯作者：
Butler, WT
中科院分区：
文献类型：
--
作者：
Qin, CL;Huang, BZ;Butler, WT
牙本质基质蛋白1（Dentin matrix protein 1，DMP 1）是一种酸性非胶原蛋白，基因消融显示其对骨和牙本质的适当矿化至关重要。在这些组织的细胞外基质中，DMP 1作为代表cDNA推导的氨基酸序列的NH 2-末端（37 kDa）和COOH-末端（57 kDa）部分的片段存在。在我们分离骨非胶原蛋白的过程中，我们观察到一种高分子量的DMP 1相关组分（命名为DMP 1-PG）。我们用抗DMP 1单克隆抗体亲和柱纯化了DMP 1-PG。氨基酸分析和胰蛋白酶肽的Edman降解证明，DMP 1-PG的核心蛋白是DMP 1的37-kDa片段。软骨素酶处理表明，DMP 1-PG的较慢的迁移速率是由于糖胺聚糖的存在。定量二糖分析表明，糖胺聚糖主要由4-硫酸软骨素组成。对胰蛋白酶肽的进一步分析使我们得出结论，单个糖胺聚糖链通过位于Ser（74）-Gly（75）二肽中的Ser（74）与核心蛋白连接，Ser（74）-Gly（75）二肽是一种对糖胺聚糖连接具有特异性的氨基酸序列。我们的研究结果表明，除了其存在的磷蛋白，从DMP 1的NH 2-末端片段发生作为蛋白聚糖。氨基酸序列比对分析表明，Ser（74）-Gly（75）二肽及其侧翼区域在从凯门鳄到智人的广泛物种中高度保守，表明该糖胺聚糖附着结构域经历了极长时间的进化压力，表明糖胺聚糖可能对DMP 1的基本生物学功能至关重要。
Dentin matrix protein 1 (DMP1) is an acidic noncollagenous protein shown by gene ablations to be critical for the proper mineralization of bone and dentin. In the extracellular matrix of these tissues DMP1 is present as fragments representing the NH2-terminal (37 kDa) and COOH-terminal (57 kDa) portions of the cDNA-deduced amino acid sequence. During our separation of bone noncollagenous proteins, we observed a high molecular weight, DMP1-related component (designated DMP1-PG). We purified DMP1-PG with a monoclonal anti-DMP1 antibody affinity column. Amino acid analysis and Edman degradation of tryptic peptides proved that the core protein for DMP1-PG is the 37-kDa fragment of DMP1. Chondroitinase treatments demonstrated that the slower migration rate of DMP1-PG is due to the presence of glycosaminoglycan. Quantitative disaccharide analysis indicated that the glycosaminoglycan is made predominantly of chondroitin 4-sulfate. Further analysis on tryptic peptides led us to conclude that a single glycosaminoglycan chain is linked to the core protein via Ser(74), located in the Ser(74)-Gly(75) dipeptide, an amino acid sequence specific for the attachment of glycosaminoglycans. Our findings show that in addition to its existence as a phosphoprotein, the NH2-terminal fragment from DMP1 occurs as a proteoglycan. Amino acid sequence alignment analysis showed that the Ser(74)-Gly(75) dipeptide and its flanking regions are highly conserved among a wide range of species from caiman to the Homo sapiens, indicating that this glycosaminoglycan attachment domain has survived an extremely long period of evolution pressure, suggesting that the glycosaminoglycan may be critical for the basic biological functions of DMP1.