Tu C P, Matsushima A, Li N Q, Rhoads D M, Srikumar K, Reddy A P, Reddy C C
J Biol Chem. 1986 Jul 15;261(20):9540-5.
The 13 forms of human liver glutathione S-transferases (GST) (Vander Jagt, D. L., Hunsaker, L. A., Garcia, K. B., and Royer, R. E. (1985) J. Biol. Chem. 260, 11603-11610) are composed of subunits in two electrophoretic mobility groups: Mr = 26,000 (Ha) and Mr = 27,500 (Hb). Preparations purified from the S-hexyl GSH-linked Sepharose 4B affinity column revealed three additional peptides at Mr = 30,800, Mr = 31,200, and Mr = 32,200. Immunoprecipitation of human liver poly(A) RNAs in vitro translation products revealed three classes of GST subunits and related peptides at Mr = 26,000, Mr = 27,500, and Mr = 31,000. The Mr = 26,000 species (Ha) can be precipitated with antisera against a variety of rat liver GSTs containing Ya, Yb, and Yc subunits, whereas the Mr = 27,500 species (Hb) can be immunoprecipitated most efficiently by antiserum against the anionic isozymes as well as a second Yb-containing isozyme (peak V) from the rat liver. The Mr = 31,000 band can be immunoprecipitated by antisera preparations against sheep liver, rat liver, and rat testis isozymes. Human liver GSTs do not have any subunits of the rat liver Yc mobility. Antiserum against the human liver GSTs did not cross-react with the Yc subunits of rat livers or brains in immunoblotting experiments. The human liver GST cDNA clone, pGTH1, selected human liver poly(A) RNAs for the Ha subunit(s) in the hybrid-selected in vitro translation experiments. Southern blot hybridization results revealed cross-hybridization of pGTH1 with the Ya, Yb, and Yc subunit cDNA clones of rat liver GSTs. This sequence homology was substantiated further in that immobilized pGTH1 DNA selected rat liver poly(A) RNAs for the Ya, Yb, and Yc subunits with different efficiency as assayed by in vitro translation and immunoprecipitation. Therefore, we have demonstrated convincingly that sequence homology as well as immunological cross-reactivity exist between GST subunits from several rat tissues and the human liver. Also, the multiple forms of human liver GSTs are most likely encoded by a minimum of three different classes of mRNAs. These results suggest a genetic basis for the subunit heterogeneity of human liver GSTs.
人类肝脏谷胱甘肽S-转移酶（GST）的13种形式（范德·贾格特，D.L.，洪萨克，L.A.，加西亚，K.B.，和罗耶，R.E.（1985年）《生物化学杂志》260，11603 - 11610）由两个电泳迁移率组的亚基组成：Mr = 26,000（Ha）和Mr = 27,500（Hb）。从S-己基谷胱甘肽连接的琼脂糖4B亲和柱上纯化的制剂显示在Mr = 30,800、Mr = 31,200和Mr = 32,200处还有另外三种肽。对人肝poly(A) RNA体外翻译产物的免疫沉淀显示出三类GST亚基及相关肽，其Mr分别为26,000、27,500和31,000。Mr = 26,000的种类（Ha）可以用针对多种含有Ya、Yb和Yc亚基的大鼠肝脏GST的抗血清沉淀，而Mr = 27,500的种类（Hb）可以被针对大鼠肝脏阴离子同工酶以及第二种含Yb的同工酶（峰V）的抗血清最有效地免疫沉淀。Mr = 31,000的条带可以被针对绵羊肝脏、大鼠肝脏和大鼠睾丸同工酶的抗血清制剂免疫沉淀。人肝脏GST没有大鼠肝脏Yc迁移率的任何亚基。在免疫印迹实验中，针对人肝脏GST的抗血清与大鼠肝脏或大脑的Yc亚基没有交叉反应。人肝脏GST cDNA克隆pGTH1在杂交选择的体外翻译实验中选择人肝poly(A) RNA用于Ha亚基。Southern印迹杂交结果显示pGTH1与大鼠肝脏GST的Ya、Yb和Yc亚基cDNA克隆有交叉杂交。通过体外翻译和免疫沉淀测定，固定化的pGTH1 DNA以不同效率选择大鼠肝脏poly(A) RNA用于Ya、Yb和Yc亚基，进一步证实了这种序列同源性。因此我们令人信服地证明了几种大鼠组织的GST亚基与人肝脏之间存在序列同源性以及免疫交叉反应性。而且，人肝脏GST的多种形式很可能由至少三类不同的mRNA编码。这些结果提示了人肝脏GST亚基异质性的遗传基础。
