Identification of interacting molecules with α-synuclein and of their function in the pathology of neurodegenerative diseases

鉴定与 α-突触核蛋白相互作用的分子及其在神经退行性疾病病理学中的功能

• 批准号: 11670960
• 负责人: NAKAI Toshiki
• 金额: $ 1.98万
• 依托单位: Yokohama City University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11670960/
• 关键词: α-synuclein; Parkinson's disease; diffuse Lewy body disease; Lewy body; synphilin; synuclein; GST fusion protein; interacting molecule; びまん性レビー小体病; レビー小体; ユビキリン; Lewy小体

α-Synuclein is regarded as one of the key molecules when considering the pathogenic mechanisms of a group of Lewy body diseases (LBD), including Parkinson's disease (PD) and diffuse Lewy body disease, in that, first, some mutations of this protein cause familial PD and, second, it is a main proteineous component of the Lewy bodies. Identification and characterization of molecules interacting with this protein is indispensable for the elucidation of the pathogenic mechanisms of LBD.This project focused on the following two points ; first, identification of new molecules interacting with α-synuclein, and second, detailed analyses of the interaction between synuclein and the known interacting molecules. In regard to the first point, we screened a human fetal brain cDNA library using A53T PD-pathogenic mutant synuclein as a bait and obtained some candidate clones, characterization of which is now under way. In addition, by probing metabolically radioisotope-labeled COS-1 proteins with GST-synucleins, we identified a polypeptide with the apparent molecular weight of 72 kDa. which interacts with GST-α-synuclein but not with GST alone. Concerning the latter point, we mainly concentrated on synphilin, which was reported during the period of our project as a protein interacting with α-synuclein. In contradiction to the previously reported results, our results indicate that, as compared with the interaction of an N- and C-terminally truncated synphilin with synuclein, that of full-length wild type one is very week if any, suggesting that presence of either of the N-terminal or C-terminal region acts inhibitory on the interaction.

在考虑包括帕金森病（PD）和弥漫性路易体病在内的一组路易体疾病（LBD）的致病机制时，α-突触核蛋白被认为是关键分子之一，因为首先，该蛋白质的一些突变会导致家族性PD，其次，它是路易体的主要蛋白质成分。与该蛋白相互作用的分子的鉴定和表征对于阐明LBD的致病机制是必不可少的。本项目重点关注以下两点：首先，鉴定与 α-突触核蛋白相互作用的新分子，其次，详细分析突触核蛋白与已知相互作用分子之间的相互作用。关于第一点，我们以A53T PD致病突变突触核蛋白为诱饵，筛选了人胎脑cDNA文库，获得了一些候选克隆，目前正在对其进行表征。此外，通过用 GST-突触核蛋白探测代谢放射性同位素标记的 COS-1 蛋白，我们鉴定出表观分子量为 72 kDa 的多肽。它与 GST-α-突触核蛋白相互作用，但不单独与 GST 相互作用。关于后一点，我们主要关注突触蛋白，在我们的项目期间，它被报道为与 α-​​突触核蛋白相互作用的蛋白质。与之前报道的结果相反，我们的结果表明，与 N 端和 C 端截短的突触蛋白与突触核蛋白的相互作用相比，全长野生型 1 的相互作用非常弱（如果有的话），这表明 N 端或 C 端区域的存在对相互作用起到抑制作用。