Recombinant Neural Protein PrP Can Bind with Both Recombinant and Native Apolipoprotein E In Vitro  

作　　者：Chen GAO Yan-Jun LEI Jun HAN Qi SHI Lan CHEN Yan GUO Yong-Jun GAO Jian-Ming CHEN Hui-Ying JIANG Wei ZHOU Xiao-Ping DONG 

机构地区：[1]State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Viral Disease Control and Prevention Chinese Center for Disease Control and Prevention, Beijing 100052, China [2]School of Medicine, Xi'an Jiaotong University, Xi'an 710061, China [3]National Laboratory of Medical Molecular Biology, Institute of Basic Medical Science, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100005, China [4]College of Science and Veterinary Medicine, Northwest Agriculture and Forest University, Yangling 712100, China

出　　处：《Acta Biochimica et Biophysica Sinica》2006年第9期593-601,共9页生物化学与生物物理学报（英文版）

基　　金：supported by the grants from the National Natural Science Foundation of China(30130070,30571672 and 30500018);the National High Technology Research and Development Program of China(2001AA215391);the National Science and Technology Task Force Project(2003BA712A04-02);the EU Project(QLRT 2000 01441).

摘　　要：The most essential and crucial step during the pathogenesis of transmissible spongiform encephalopathy is the conformational change of cellular prion protein （PrP^C） to pathologic isoform （prp^Sc）. A lot of data revealed that caveolae-like domains （CLDs） in the cell surface were the probable place where the conversion of PrP proteins happened. Apolipoprotein E （ApoE） is an apolipoprotein which is considered to play an important role in the development of Alzheimer＇s disease and other neurodegenerative diseases by forming protein complex through binding to the receptor located in the clathrin-coated pits of the cell surface. In this study, a 914-bp cDNA sequence encoding human ApoE3 was amplified from neuroblastoma cell line SH-SY5Y. Three human ApoE isomers were expressed and purified from Escherichia coli. ApoE-specific antiserum was prepared by immunizing rabbits with the purified ApoE3. GST/His pull-down assay, immunoprecipitation and ELISA revealed that three full-length ApoE isomers interact with the recombinant full-length PrP protein in vitro. The regions corresponding to protein binding were mapped in the N-terminal segment of ApoE （amino acid 1-194） and the N-terminal of PrP （amino acid 23-90）. Moreover, the recombinant PrP showed the ability to form a complex with the native ApoE from liver tissues. Our data provided direct evidence of molecular interaction between ApoE and PrP. It also supplied scientific clues for assessing the significance of CLDs on the surface of cellular membrane in the process of conformational conversion from PrP^C to PrP^Sc and probing into the pathogenesis of transmissible spongiform encephalopathy.The most essential and crucial step during the pathogenesis of transmissible spongiform encephalopathy is the conformational change of cellular prion protein （PrP^C） to pathologic isoform （prp^Sc）. A lot of data revealed that caveolae-like domains （CLDs） in the cell surface were the probable place where the conversion of PrP proteins happened. Apolipoprotein E （ApoE） is an apolipoprotein which is considered to play an important role in the development of Alzheimer＇s disease and other neurodegenerative diseases by forming protein complex through binding to the receptor located in the clathrin-coated pits of the cell surface. In this study, a 914-bp cDNA sequence encoding human ApoE3 was amplified from neuroblastoma cell line SH-SY5Y. Three human ApoE isomers were expressed and purified from Escherichia coli. ApoE-specific antiserum was prepared by immunizing rabbits with the purified ApoE3. GST/His pull-down assay, immunoprecipitation and ELISA revealed that three full-length ApoE isomers interact with the recombinant full-length PrP protein in vitro. The regions corresponding to protein binding were mapped in the N-terminal segment of ApoE （amino acid 1-194） and the N-terminal of PrP （amino acid 23-90）. Moreover, the recombinant PrP showed the ability to form a complex with the native ApoE from liver tissues. Our data provided direct evidence of molecular interaction between ApoE and PrP. It also supplied scientific clues for assessing the significance of CLDs on the surface of cellular membrane in the process of conformational conversion from PrP^C to PrP^Sc and probing into the pathogenesis of transmissible spongiform encephalopathy.

关 键 词：prion disease PRP apolipoprotein E protein interaction caveolae-like domain 

分 类 号：Q51[生物学—生物化学]