Prokaryotic Expression,Purification,Antibody Production of a NH_2-Terminal Fragment of mCLCA3 Protein and Analysis of mClCA3 Protein Expression in Asthmatic Mouse Lung  

作　　者：HOU Xia WU Qing-tian ZHANG Yu-ping ZHU Na LIU Yan-li FENG Xue-chao MA Tong-hui 

机构地区：[1]Membrane Channel Research Laboratory, Northeast Normal University, Changchun 130024, P. R. China [2]School of Basic Medicine, Jiamusi University, Jiamusi 154002, P. R. China

出　　处：《Chemical Research in Chinese Universities》2007年第6期688-692,共5页高等学校化学研究（英文版）

基　　金：Supported by the Distinguished Young Scholars Fund of China(No.30325011);the National Natural Science Foundation of China(Nos.30470405and30670477);Distinguished Young Scholars Fund of Jilin Province,China(No.20030112);Excellent Young Teachers Program of MOE,China.

摘　　要：mCLCA3 is a member of calcium activated chloride channel（CACC） family that may play an important role in mucin packaging and secretion in asthmatic and cystic fibrosis lung. To study the protein structure and expression of mCLCA3 in asthmatic mouse lung, an N-terminal 269 amino acid peptide of mCLCA3 was expressed in E. coli, purified to homogeneity and rabbit polyclonal antibodies against this peptide were generated. Immunohistochemistry of asthmatic mouse lung using the antibody indicated exclusive mCLCA3 expression in mucin granules of goblet cells in airway surface and lumen, Immunoblot analysis of lavage fluid from asthmatic mouse lung revealed a single 90 kDa protein form of mClCA3. The results demonstrate that the 90 kDa N-terminal peptide, neither the flail-length protein nor the reported N-terminal 35 kDa cleaved form of mClCA3 is the major functional form involved in the packaging and exocytosis of mucin granules in asthmatic goblet cells.mCLCA3 is a member of calcium activated chloride channel（CACC） family that may play an important role in mucin packaging and secretion in asthmatic and cystic fibrosis lung. To study the protein structure and expression of mCLCA3 in asthmatic mouse lung, an N-terminal 269 amino acid peptide of mCLCA3 was expressed in E. coli, purified to homogeneity and rabbit polyclonal antibodies against this peptide were generated. Immunohistochemistry of asthmatic mouse lung using the antibody indicated exclusive mCLCA3 expression in mucin granules of goblet cells in airway surface and lumen, Immunoblot analysis of lavage fluid from asthmatic mouse lung revealed a single 90 kDa protein form of mClCA3. The results demonstrate that the 90 kDa N-terminal peptide, neither the flail-length protein nor the reported N-terminal 35 kDa cleaved form of mClCA3 is the major functional form involved in the packaging and exocytosis of mucin granules in asthmatic goblet cells.

关 键 词：mCLCA3 Prokaryotic expression PURIFICATION Antibody production ASTHMA 

分 类 号：Q78[生物学—分子生物学]