New mammalian selenocysteine-containing proteins identified with an algorithm that searches for selenocysteine insertion sequence elements
New mammalian selenocysteine-containing proteins identified with an algorithm that searches for selenocysteine insertion sequence elements
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DOI：
10.1074/jbc.274.48.33888
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发表时间：
1999-11-26
影响因子：
4.8
通讯作者：
Gladyshev, VN
中科院分区：
文献类型：
--
作者：
Kryukov, GV;Kryukov, VM;Gladyshev, VN
目前鉴定的哺乳动物含硒蛋白含有由UGA编码的硒代半胱氨酸残基形式的硒，这些蛋白缺乏共同的氨基酸序列基序，但硒蛋白基因的3 '-非翻译区含有共同的茎环结构，硒代半胱氨酸插入序列（SECTS）元件，其是将UGA解码为硒代半胱氨酸所必需的，而不是终止信号。本文介绍了一个计算机程序SECISTORM，它根据SECIS元件的一级结构、二级结构和自由能要求，识别哺乳动物硒蛋白基因。当SECISTORM应用于寻找人类dbEST时，发现了两个新的哺乳动物硒蛋白，命名为SelT和SelR。我们确定了它们的cDNA序列，并在猴细胞系中将其表达为与绿色荧光蛋白的融合蛋白。硒-75代谢标记证实了硒纳入新的蛋白质，并在免疫印迹试验中另外记录了SelT的表达，SelT和SelR与先前表征的蛋白质没有同源性，但在各种生物体中检测到它们推定的同源物。SelR同源基因存在于每一个生物体的全基因组测序的特点，数据表明SECISTORM鉴定新的硒蛋白基因的核苷酸数据库的适用性。
Mammalian selenium-containing proteins identified thus far contain selenium in the form of a selenocysteine residue encoded by UGA, These proteins lack common amino acid sequence motifs, but 3'-untranslated regions of selenoprotein genes contain a common stem-loop structure, selenocysteine insertion sequence (SECTS) element, that is necessary for decoding UGA as selenocysteine rather than a stop signal. We describe here a computer program, SECISearch, that identifies mammalian selenoprotein genes by recognizing SECIS elements on the basis of their primary and secondary structures and free energy requirements, When SECISearch was applied to search human dbEST, two new mammalian selenoproteins, designated SelT and SelR, were identified. We determined their cDNA sequences and expressed them in a monkey cell line as fusion proteins with a green fluorescent protein. Incorporation of selenium into new proteins was confirmed by metabolic labeling with Se-75, and expression of SelT was additionally documented in immunoblot assays, SelT and SelR did not have homology to previously characterized proteins, but their putative homologs were detected in various organisms. SelR homologs were present in every organism characterized by complete genome sequencing, The data suggest applicability of SECISearch for identification of new selenoprotein genes in nucleotide data bases.