Identification and structural and functional characterization of human enamelysin (MMP-20)
Identification and structural and functional characterization of human enamelysin (MMP-20)
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DOI：
10.1021/bi972120y
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发表时间：
1997-12-09
期刊：
影响因子：
2.9
通讯作者：
LopezOtin, C
中科院分区：
文献类型：
--
作者：
Llano, E;Pendas, AM;LopezOtin, C
从成牙细胞制备的RNA中克隆出一种新的人基质金属蛋白酶（MMP） cDNA。克隆的cDNA的开放阅读框编码一个483个氨基酸的多肽，并且与最近描述的猪漆膜酵素的序列广泛相似，表明分离的cDNA编码该酶的人类同源物。Human enamelysin （MMP-20）具有与其他MMPs类似的结构域组织，包括一个信号肽，一个具有保守基序PRCGVPD的前结构域，一个具有锌结合位点的催化结构域，以及一个类似于血凝素序列的cooh末端片段。人釉溶素的计算分子量约为54 kDa，与胶原酶或基质溶素相似。然而，这种人类MMP缺乏这些MMP亚家族特有的一系列结构特征。已在大肠杆菌中表达了全长的人搪瓷酶cDNA，纯化后的重组蛋白能够降解作为MMPs底物的合成肽，证实了人搪瓷酶属于该蛋白酶家族。此外，重组人釉质酶能够降解釉质基质的主要蛋白质成分——淀粉原蛋白。基于其对淀粉原蛋白的降解活性，以及其在牙组织中的高度限制性表达，我们认为人釉质酶在牙釉质形成过程中起着核心作用。最后，我们发现人类釉溶素基因（MMP-20）位于染色体11q22上，与MMP基因家族的至少7个其他成员聚集在一起。
A cDNA encoding a new human matrix metalloproteinase (MMP) has been cloned from RNA prepared from odontoblastic cells. The open reading frame of the cloned cDNA codes for a polypeptide of 483 amino acids and is extensively similar to the sequence of recently described porcine enamelysin, suggesting that the isolated cDNA codes for the human homologue of this enzyme. Human enamelysin (MMP-20) has a domain organization similar to other MMPs, including a signal peptide, a prodomain with the conserved motif PRCGVPD involved in maintaining enzyme latency, a catalytic domain with a Zn-binding site, and a COOH-terminal fragment similar to the sequence of hemopexin. The calculated molecular mass of human enamelysin is about 54 kDa, which is similar to that of collagenases or stromelysins. However, this human MMP lacks a series of structural features distinctive of these subfamilies of MMPs. The full-length human enamelysin cDNA has been expressed in Escherichia coli, and the purified and refolded recombinant protein is able to degrade synthetic peptides used as substrates of MMPs, confirming that human enamelysin belongs to this family of proteases. Furthermore, the recombinant human enamelysin is able to degrade amelogenin, the major protein component of the enamel matrix. On the basis of its degrading activity on amelogenin, and its highly restricted expression to dental tissues, we suggest that human enamelysin plays a central role in the process of tooth enamel formation. Finally, we have found that the human enamelysin gene (MMP-20) maps to chromosome 11q22, clustered to at least seven other members of the MMP gene family.