Elucidation of characters and neurotoxic mechanisms of alpha-synuclein oligomers and its application to molecular targeted therapy

我们建立了制备α-突触核蛋白(A-SYN)低聚体和原纤维的方法，即在37℃下培养高浓度的重组A-突触核蛋白(A-SYN)低聚体和原纤维7天。虽然重组A-SYN分子和来自人脑脊液的天然A-SYN分子在60-70 kDa左右的凝胶过滤层析中得到洗脱，但这些分子不能与8M尿素解离，因此已被证明在水溶液中以单体形式存在。在培养细胞中，重组A-syn寡聚体和原纤维的混合物可形成细胞内聚集体，其降解依赖于自噬途径。

We established the preparing method of alpha-synuclein (A-syn) oligomers and protofibrils by incubating high-concentration solution, 3 mg/mL, of recombinant A-syn with stirring for 7 days at 37 C. Although recombinant A-syn molecules and natural A-syn molecules derived from human CSF were eluted from gel-filtration chromatography at around fractions of 60-70 kDa, those molecules were not dissociated with 8M urea, and accordingly have been demonstrated to exist as monomers in aqueous solution. In cultured cells, intracellular aggregates were formed by adding mixture of recombinant A-syn oligomers and protofibrils into the culture medium, and their degradation was dependent on the autophagy pathway.