中文摘要：

目的：确定α-Synuclein蛋白与线粒体相互作用的功能结构域,并检测该结构域对线粒体功能的影响。方法：构建重组融合蛋白质粒pCMV-myc/α-Syn-WT、pCMV-myc/α-Syn-N和pCMV-myc/α-Syn-△N,转染人胚肾HEK293T细胞,通过免疫共沉淀明确α-Synuclein蛋白与线粒体相互作用的功能结构域。使用表达α-Synuclein蛋白的病毒上清感染小鼠多巴胺能神经细胞MN9D,通过免疫荧光、流式细胞术检测线粒体膜电位及Cytochrome c释放。结果：成功构建了pCMV-myc/α-Syn-WT、pCMV-myc/α-Syn-N和pCMV-myc/α-Syn-△N融合蛋白质粒,免疫共沉淀明确α-Synuclein N-端为其功能结构域,JC-1染色发现N-端使线粒体膜电位降低,流式细胞术证实N-端使Cytochrome c释放明显增加。结论：α-Synuclein N-端是其与线粒体相互作用的功能结构域,N-端降低线粒体功能。

英文摘要：

Objective： To identify the functional domain of α-Synuclein in affecting mitochondrial function and how the function to be impaired, especially, the mitochondrial membrane potential and the release of Cytochrome e. Methods： Harvest of α-Syn-N and α-Syn-△N by PCR, then subcloned into the pCMV-Myc mammalian expression vector. The recombinant plasmids were transfected into HEK293T cells by Lipofectamine 2000. After detecting the protein expression by Western blot, the functional domain was detected by coimmunoprecipitation. The mitochondrial membrane potential through flow cytometry and immunofluorescence, at the same time, the release of Cytochrome c through flow cytometry to detect. Results： The recombinant plasmids were constructed successfully. CO-IP has proved that N-terminal may be the functional domain of α-Synuclein in affecting mitochondria. Over-expression of N-terminal could depolarize the mitochondrial membrane potential and induce the Cytochrome c releasing in MN9D cells. Conclusion： N-terminal may be the functional domain of α- synuclein and over-expression of N-terminal could decrease mitochondrial activity.