Rat liver glutathione S-transferases. Construction of a cDNA clone complementary to a Yc mRNA and prediction of the complete amino acid sequence of a Yc subunit.
Rat liver glutathione S-transferases. Construction of a cDNA clone complementary to a Yc mRNA and prediction of the complete amino acid sequence of a Yc subunit.
复制标题
大鼠肝脏谷胱甘肽 S-转移酶。
DOI：
10.1016/s0021-9258(18)89095-0
复制
发表时间：
1985
期刊：
影响因子：
--
通讯作者：
Cecil B. PickettS
中科院分区：
文献类型：
--
作者：
Claudia A. Telakowski;J. Rodkey;Carl D. Bennett;Anthony Y. H. Lull;Cecil B. PickettS
如Gubler和霍夫曼（Gubler，U.，和霍夫曼，B. S.（1983）Gene 25，263-269）。重组克隆pGTB 42含有900个碱基对的cDNA插入片段，其3'端在杂交选择翻译实验中显示出对Yc mRNA的特异性。测定了pGTB 42的核苷酸序列，并推导出Yc亚基的完整氨基酸序列。该cDNA克隆含有663个核苷酸的开放阅读框，编码包含221个氨基酸的多肽，分子量为25，322。从pGTB 42推导的NH 2-末端序列与通过常规蛋白质测序技术确定的Ya-Yc异源二聚体的前39个氨基酸一致。pGTB 42的核苷酸序列与我们实验室先前描述的Ya克隆pGTB 38的序列的比较（Pickett，C. B.，Telakowski-Hopkins，C.一、丁氏G. J. - F.、科布莱特湖，和Lu，A.Y.H.（1984）J.Biol.Chem.259，5182-5188）揭示了在两个克隆的相同区域上66%的序列同源性;然而，Ya和Yc mRNA的5 ′-和3 ′-非翻译区在它们的序列上是完全不同的。Ya和Yc亚基之间的总体氨基酸序列同源性为68%，然而，NH 2-末端结构域比中间或羧基-末端结构域更高度保守。我们的数据表明，大鼠肝脏谷胱甘肽S-转移酶的Ya和Yc亚基是来自两个相关但不同基因的两种不同mRNA的产物。
Using polysomal immunoselected rat liver glutathione S-transferase mRNAs, we have constructed cDNA clones using DNA polymerase I, RNase H, and Escherichia coli ligase (NAD+)-mediated second strand cDNA synthesis as described by Gubler and Hoffman (Gubler, U., and Hoffman, B. S. (1983) Gene 25, 263-269). Recombinant clone, pGTB42, contained a cDNA insert of 900 base pairs whose 3' end showed specificity for the Yc mRNA in hybrid-select translation experiments. The nucleotide sequence of pGTB42 has been determined, and the complete amino acid sequence of a Yc subunit has been deduced. The cDNA clone contains an open reading frame of 663 nucleotides encoding a polypeptide comprising 221 amino acids with a molecular weight of 25,322. The NH2-terminal sequence deduced from pGTB42 is in agreement with the first 39 amino acids determined for a Ya-Yc heterodimer by conventional protein-sequencing techniques. A comparison of the nucleotide sequence of pGTB42 with the sequence of a Ya clone, pGTB38, described previously by our laboratory (Pickett, C. B., Telakowski-Hopkins, C. A., Ding, G. J.-F., Argenbright, L., and Lu, A.Y.H. (1984) J. Biol. Chem. 259, 5182-5188) reveals a sequence homology of 66% over the same regions of both clones; however, the 5'- and 3'-untranslated regions of the Ya and Yc mRNAs are totally divergent in their sequences. The overall amino acid sequence homology between the Ya and Yc subunits is 68%, however, the NH2-terminal domain is more highly conserved than the middle or carboxyl-terminal domains. Our data suggest that the Ya and Yc subunits of the rat liver glutathione S-transferases are products of two different mRNAs which are derived from two related yet different genes.