编码翻译起始因子6的人cDNA的分子克隆与功能表达
Molecular cloning and functional expression of a human cDNA encoding translation initiation factor 6.
作者信息
Si K, Chaudhuri J, Chevesich J, Maitra U
机构信息
Department of Developmental and Molecular Biology, Albert Einstein College of Medicine of Yeshiva University, Jack and Pearl Resnick Campus, Bronx, NY 10461, USA.
出版信息
Proc Natl Acad Sci U S A. 1997 Dec 23;94(26):14285-90. doi: 10.1073/pnas.94.26.14285.
Abstract
Eukaryotic translation initiation factor 6 (eIF6) binds to the 60S ribosomal subunit and prevents its association with the 40S ribosomal subunit. In this paper, we devised a procedure for purifying eIF6 from rabbit reticulocyte lysates and immunochemically characterized the protein by using antibodies isolated from egg yolks of laying hens immunized with rabbit eIF6. By using these monospecific antibodies, a 1.096-kb human cDNA that encodes an eIF6 of 245 amino acids (calculated Mr 26,558) has been cloned and expressed in Escherichia coli. The purified recombinant human protein exhibits biochemical properties that are similar to eIF6 isolated from mammalian cell extracts. Database searches identified amino acid sequences from Saccharomyces cerevisiae, Drosophila, and the nematode Caenorhabditis elegans with significant identity to the deduced amino acid sequence of human eIF6, suggesting the presence of homologues of human eIF6 in these organisms.
摘要
真核生物翻译起始因子6（eIF6）与60S核糖体亚基结合，并阻止其与40S核糖体亚基结合。在本文中，我们设计了一种从兔网织红细胞裂解物中纯化eIF6的方法，并使用从用兔eIF6免疫的产蛋母鸡蛋黄中分离的抗体对该蛋白质进行了免疫化学表征。通过使用这些单特异性抗体，克隆了一个编码245个氨基酸（计算分子量为26,558）的eIF6的1.096 kb人cDNA，并在大肠杆菌中进行了表达。纯化的重组人蛋白表现出与从哺乳动物细胞提取物中分离的eIF6相似的生化特性。数据库搜索鉴定出酿酒酵母、果蝇和线虫秀丽隐杆线虫的氨基酸序列与推导的人eIF6氨基酸序列具有显著同源性，表明这些生物体中存在人eIF6的同源物。
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