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仓鼠和鸡细胞中高度保守的葡萄糖调节蛋白:其cDNA克隆的初步表征

Highly conserved glucose-regulated protein in hamster and chicken cells: preliminary characterization of its cDNA clone.

作者信息

Lee A S, Delegeane A, Scharff D

出版信息

Proc Natl Acad Sci U S A. 1981 Aug;78(8):4922-5. doi: 10.1073/pnas.78.8.4922.

Abstract

A temperature-sensitive mutant K12 derived from a Chinese hamster fibroblast has been shown to overproduce three specific proteins of Mr 94,000, 78,000, and 58,000 when incubated at the nonpermissive temperature (40.5 degrees C). We previously identified these proteins as glucose-regulated proteins similar to those observed in chicken embryo fibroblasts when the cells are starved of glucose. In this report, we show that the Mr 78,000 proteins isolated from the hamster K12 cell line and from chicken embryo fibroblasts have identical electrophoretic mobilities in two-dimensional isoelectric focusing gels and nearly identical peptide maps. However, these proteins are different from heat-shock proteins previously described for animal cells. We have constructed a library of cDNA clones by using the RNA extracted from the hamster K12 cells incubated at 40.5 degrees C. Clones that hybridize preferentially with cDNA made from RNA at 40.5 degrees C were selected. By using the hybrid-selection technique, followed by in vitro translation, a cDNA clone containing a 2550-nucleotide insert coding for the hamster Mr 78,000 protein has been identified.

摘要

源自中国仓鼠成纤维细胞的温度敏感突变体K12已被证明,在非允许温度(40.5摄氏度)下孵育时会过量产生三种特定蛋白质,其分子量分别为94,000、78,000和58,000。我们之前将这些蛋白质鉴定为葡萄糖调节蛋白,类似于鸡胚成纤维细胞在缺乏葡萄糖时所观察到的那些蛋白。在本报告中,我们表明,从仓鼠K12细胞系和鸡胚成纤维细胞中分离出的分子量为78,000的蛋白质,在二维等电聚焦凝胶中具有相同的电泳迁移率,并且肽图几乎相同。然而,这些蛋白质与先前描述的动物细胞热休克蛋白不同。我们使用从在40.5摄氏度下孵育的仓鼠K12细胞中提取的RNA构建了一个cDNA克隆文库。选择了优先与在40.5摄氏度下由RNA制成的cDNA杂交的克隆。通过使用杂交选择技术,随后进行体外翻译,已鉴定出一个含有2550个核苷酸插入片段的cDNA克隆,该片段编码仓鼠分子量为78,000的蛋白质。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/690f/320295/1adb6de7e68c/pnas00659-0298-a.jpg

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