A Preliminary Study Of The Expression Of GABA Receptor Subcloned Three Organisms

Type A γ-aminobutyric acid (aminobutyric acid, GABA) receptor, which is a classof ligand-gated ion channel receptor, is the main inhibitory neurotransmitter in thecentral nervous system of vertebrates and non-vertebrates. The receptor is made upof homopentamer or heterpentamer. There are many drug binding sites on theGABAA receptor, so it became a great hotspot for pharmaceutical and pesticideresearchers. However, the quantities of these natural receptors are low. The cloneand expression of Drosophila GABA receptor, which has bioactivity, presentsimportant references for the development of new, efficient and safe pesticide. In thispaper, we attempt to attain part of the subclone and heterologous expression of thevertebrates and insect GABA mutant receptor anti-dieldrin (RDL) by the molecularsubcloning technique.In this experiment, we used the information technology of molecular biology toobtain the information of the overall and partial structure of Drosophila GABAreceptor. Searching from the NCBI library, the pET-21a was used to build theexpression vector with the full length and truncated part of the N-terminalextracellular region of the whole Drosophila RDL sequence. At the conditions of37℃, IPTG0.5mM, we induced fusion strain, which is the E.coli BL21(DE3) cells, for3h. It can express full-length protein containing the HisTag. However, the truncatedportion didnt expressed successfully. Detected by SDS-PAGE electrophoresis andwestern blot, the specificity of the expressed protein is correct, but the purity andamount are not enough for the follow-up test.It is the first paper that try to express the full-length of the eukaryotic animalGABA receptor (RDL) in the prokaryotic system.