Sequential Extraction of Soluble and Insoluble Alpha-Synuclein from Parkinsonian Brains.
Sequential Extraction of Soluble and Insoluble Alpha-Synuclein from Parkinsonian Brains.
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DOI：
10.3791/53415
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发表时间：
2016-01-05
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影响因子：
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通讯作者：
Bandopadhyay R
中科院分区：
文献类型：
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作者：
Bandopadhyay R
α-突触核蛋白（α-syn）蛋白主要在神经元内大量表达，并且以许多不同的形式存在-单体、四聚体、寡聚体和原纤维。在疾病期间，α-syn经历构象变化以形成寡聚体和高分子量聚集体，其倾向于使蛋白质更难溶解。异常聚集的α-syn是帕金森病（PD）、路易体痴呆（DLB）和多系统萎缩（MSA）的神经病理学特征。使用具有增加的去污剂强度和高速超离心的缓冲液对不溶性α-syn进行生化表征和分析，为确定与疾病进展相关的α-syn病理学的发展提供了有力的工具。该方案描述了从死后人脑组织中分离越来越不溶性/聚集的α-syn。这种方法可以通过修改适用于在含有不同α-syn突变的转基因动物模型中以及在其他神经退行性疾病中研究正常和异常α-syn生物学，这些疾病的特征是与其各自病理相关的蛋白质的异常纤维沉积。
Alpha-synuclein (α-syn) protein is abundantly expressed mainly within neurons, and exists in a number of different forms - monomers, tetramers, oligomers and fibrils. During disease, α-syn undergoes conformational changes to form oligomers and high molecular weight aggregates that tend to make the protein more insoluble. Abnormally aggregated α-syn is a neuropathological feature of Parkinson's disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). Biochemical characterization and analysis of insoluble α-syn using buffers with increasing detergent strength and high-speed ultracentrifugation provides a powerful tool to determine the development of α-syn pathology associated with disease progression. This protocol describes the isolation of increasingly insoluble/aggregated α-syn from post-mortem human brain tissue. This methodology can be adapted with modifications to studies of normal and abnormal α-syn biology in transgenic animal models harbouring different α-syn mutations as well as in other neurodegenerative diseases that feature aberrant fibrillar deposits of proteins related to their respective pathologies.