Sequencing of bovine enamelin ("tuftelin") a novel acidic enamel protein.
Sequencing of bovine enamelin ("tuftelin") a novel acidic enamel protein.
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DOI：
10.1016/s0021-9258(18)98510-8
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发表时间：
1991-08
期刊：
影响因子：
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通讯作者：
D. Deutsch;Aron Palmon;Larry W. Fisherll;N. Kolodny;John D. Terminell;Marian F. Youngll
中科院分区：
文献类型：
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作者：
D. Deutsch;Aron Palmon;Larry W. Fisherll;N. Kolodny;John D. Terminell;Marian F. Youngll
釉质是一组主要的28-70-kDa的酸性蛋白质，富含天冬氨酸、谷氨酸、丝氨酸和甘氨酸，存在于发育和成熟的细胞外釉质中;是覆盖脊椎动物牙齿的独特且高度矿化的外胚层组织。它们与该组织的矿化和结构组织有关。为了阐明釉蛋白的一级结构，对从牛成釉细胞富集的λ Zap 2表达文库中分离的2674个碱基对的cDNA进行了测序。通过氨基酸组成、酶联免疫吸附试验、Western blotting、间接免疫组化和高分辨率蛋白质-A金免疫细胞化学证实了推导蛋白的身份和定位。使用针对对应于从克隆的cDNA序列推导的蛋白质序列的合成肽的抗体，采用免疫学技术。结果表明，该蛋白是一种新的酸性釉蛋白。它含有389个氨基酸，计算分子量为43，814。其氨基酸组成类似于“簇”蛋白（残留在成熟组织中的釉质基质蛋白片段）。它含有一个潜在的N-糖基化位点和5个半胱氨酸残基。克隆的cDNA与牛基因组DNA的Southern杂交表明，存在一个单一的基因与一个或多个内含子。
Enamelins are a major group of 28-70-kDa acidic proteins rich in aspartic acid, glutamic acid, serine, and glycine found in developing and mature extracellular enamel; a unique and highly mineralized ectodermal tissue covering vertebrate teeth. They have been associated with the mineralization and structural organization of this tissue. In an attempt to elucidate the primary structure of enamelin, a 2674-base pair cDNA isolated from a bovine ameloblast-enriched, lambda Zap 2 expression library, was sequenced. The identity and localization of the deduced protein was confirmed by amino acid composition, enzyme-linked immunosorbent assay, Western blotting, indirect immunohistochemistry, and high resolution protein-A gold immunocytochemistry. The immunological techniques were employed using antibodies directed against synthetic peptides corresponding to the protein sequence deduced from the cloned cDNA sequence. The results reveal the deduced protein to be a novel acidic enamel protein. It contains 389 amino acids and has a calculated molecular weight of 43,814. Its amino acid composition is similar to that of “tuft” proteins (enamel matrix protein fragments remaining in the mature tissue). It contains one potential N-glycosylation site and 5 cysteine residues. Southern hybridization of the cloned cDNA with genomic bovine DNA indicated the existence of a single gene with one or more introns.