Selection,Expression and Purification of Human Abzyme Containing Selenium with Type Ⅰ Thyroxine Deiodinase Activity  

作　　者：LIN Feng LIU Lan-ying NI Jia-zuan 

机构地区：[1]College of Life Science, Jilin University, Changchun 130012, P. R. China, [2]Key Laboratory of Rare Earth of Chinese Academy, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, P. R. China [3]College of Life Science, Shenzhen University, Shenzhen 518060, P. R. China

出　　处：《Chemical Research in Chinese Universities》2011年第4期619-622,共4页高等学校化学研究（英文版）

基　　金：Supported by the Basic Science Research Fund of Jilin University,China(No.200903259)

摘　　要：Single-chain fragment variable（ScFv） antibodies with the substrate binding sites were obtained by repetitive selections from a semi-synthetic phage display antibody library against two haptens——thyroxine（T4） and O-methyl-T4（O-CH3-T4）.The positive phage clones were determined by ELISA and then cloned into vector pET30a（＋）.The recombined plasmids were identified by DNA sequence analysis and transformed into E.coli BL21（DE3）.The expressed proteins were isolated,purified,identified by Western blot analysis,and finally the catalytic group——selenocysteine was incorporated into the antibodies binding sites by chemical mutation.The type·thyroxine deiodinase activities of the abzymes were 0.006 and 0.008 pg·mL-1·min-1,respectively,which were about one tenth that of the mouse intact antibody that was obtained from hybridoma.Single-chain fragment variable（ScFv） antibodies with the substrate binding sites were obtained by repetitive selections from a semi-synthetic phage display antibody library against two haptens——thyroxine（T4） and O-methyl-T4（O-CH3-T4）.The positive phage clones were determined by ELISA and then cloned into vector pET30a（＋）.The recombined plasmids were identified by DNA sequence analysis and transformed into E.coli BL21（DE3）.The expressed proteins were isolated,purified,identified by Western blot analysis,and finally the catalytic group——selenocysteine was incorporated into the antibodies binding sites by chemical mutation.The type·thyroxine deiodinase activities of the abzymes were 0.006 and 0.008 pg·mL-1·min-1,respectively,which were about one tenth that of the mouse intact antibody that was obtained from hybridoma.

关 键 词：Phage antibody library HUMANIZATION SELENIUM Catalytic antibody SELECTION 

分 类 号：Q55[生物学—生物化学] Q513