Development of an anionic sensor for living cells: application to the human glycine receptor
申请人:INSTITUT PASTEUR
公开号:EP1514922A1
公开(公告)日:2005-03-16
The subject of the invention is a polypeptide comprising a new mutant enhanced yellow fluorescent protein (EYFP) having anion binding properties, which may be further fused with an enhanced cyan fluorescent protein (ECFP) and further to a peptide linker inserted between said mutant EYFP and ECFP, thereby forming an anionic sensor, said anionic sensor may further be inserted in a transmembrane anionic channel. A complex comprising a transmembrane ionic channel and a polypeptidic biosensor is also disclosed. Also disclosed are nucleic acids and vectors, host cells and transgenic organisms thereof. Also disclosed herein are a method for preparing transformed cells capable of expressing ionic sensors at the vicinity of said ionic channels, methods for determining and monitoring dynamic changes in ion concentrations in cells, methods of selecting compounds capable of modifying said dynamic changes, sensing fluorescent microspheres and kits thereof.
本发明的主题是一种多肽,包括一种具有阴离子结合特性的新突变增强型黄色荧光蛋白(EYFP),该蛋白可进一步与增强型青色荧光蛋白(ECFP)融合,并进一步与插入所述突变EYFP和ECFP之间的肽链连接,从而形成阴离子传感器,所述阴离子传感器可进一步插入跨膜阴离子通道。还公开了一种由跨膜离子通道和多肽生物传感器组成的复合物。还公开了核酸和载体、宿主细胞及其转基因生物。本文还公开了制备能在所述离子通道附近表达离子传感器的转化细胞的方法、确定和监测细胞中离子浓度动态变化的方法、选择能改变所述动态变化的化合物的方法、传感荧光微球及其试剂盒。