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目的研究支架蛋白PDLIM5与神经元α7烟碱乙酰胆碱受体(α7nAChR)间的相互作用及其作用位点。方法构建PDLIM5基因及其截短片段原核、真核重组表达载体和构建α7nAChR真核重组表达载体,并在大肠杆菌中表达与纯化相关蛋白。采用GST pull down及免疫共沉淀(CO-IP)方法检测PDLIM5与α7nAChR分子间相互作用及结合位点。结果 GST pull down和CO-IP方法检测结果显示PDLIM5与α7nAChR具有相互作用,且α7nAChR与PDLIM5中的PDZ结构域具有特异的相互作用,但不与PDLIM5中的LIM结构域发生反应。结论 PDLIM5通过其PDZ结构域介导同α7nAChR的相互作用。
Objective To investigate the interaction between the PDLIM5 scaffold protein and α7 nicotinic acetylcholine receptor (α7nAChR) in neurons and its site of action. Methods The prokaryotic and eukaryotic recombinant expression vector of PDLIM5 gene and its truncated fragment was constructed and eukaryotic recombinant expression vector of α7 nAChR was constructed and expressed and purified in E. coli. GST pull down and co-immunoprecipitation (CO-IP) were used to detect the interaction and binding sites between PDLIM5 and α7nAChR. Results The results of GST pull down assay and CO-IP assay showed that PDLIM5 interacted with α7nAChR and α7nAChR had specific interaction with PDZ domain in PDLIM5 but not with LIM domain in PDLIM5. Conclusion PDLIM5 mediates the interaction with α7 nAChR through its PDZ domain.