Cleavage of normal and pathological forms of α-synuclein by neurosin in vitro
Cleavage of normal and pathological forms of α-synuclein by neurosin in vitro
复制标题
DOI：
10.1016/j.neulet.2008.02.057
复制
发表时间：
2008-05-02
影响因子：
2.5
通讯作者：
Mizuno, Toshiki
中科院分区：
文献类型：
--
作者：
Kasai, Takashi;Tokuda, Takahiko;Mizuno, Toshiki
Neurosin 是主要在中枢神经系统中表达的丝氨酸蛋白酶之一。神经素被认为在α-突触核蛋白（α-syn）的降解中发挥重要作用，因为先前的研究表明神经素可降解α-syn，在体外抑制α-syn的聚合，并且存在于路易体中。然而，神经素降解α-syn的细节却鲜为人知。我们通过免疫印迹和液相色谱-离子阱质谱 (LC/MS/MS) 研究了神经素介导的 α-syn 裂解。我们还比较了磷酸化和非磷酸化形式的 α-syn 以及突变型和野生型 α-syn 之间神经素对 α-syn 的降解。 Neurosin 在特定位点裂解 α-syn。主要切割位点位于 NAC 区域内的 Lys80 和 Thr81 之间（E61 至 V95），这对于 α-syn 聚合很重要，因此可能会阻止 α-syn 聚合。同时，也出现了替代性的、较小形式的处理。它们通过截短 C 端区域来保留 NAC 区域，因此可能有助于 α-syn 聚合。磷酸化的 α-syn 比非磷酸化的 α-syn 更能抵抗神经素的降解。 A30P 突变体比野生型和其他 α-syn 突变体更能抵抗降解。这种对神经素介导的磷酸化 α-syn 和 A30P 突变体降解的抵抗力，分别是与帕金森病 (PD) 发展相关的翻译后因素和遗传因素，提供了神经素参与 PD 发病机制的支持证据。 (C) 2008 Elsevier Ireland Ltd. 保留所有权利。
Neurosin is one of the serine proteases predominantly expressed in the central nervous system. Neurosin is presumed to play an important role in the degradation of alpha-synuclein (alpha-syn), since a previous study showed that neurosin degrades alpha-syn, inhibits polymerization of alpha-syn in vitro, and exists in Lewy bodies. However, the details of alpha-syn degradation by neurosin are little known. We investigated neurosin-mediated cleavage of alpha-syn by immunoblotting and liquid chromatography-ion trap mass spectrometry (LC/MS/MS). We also compared alpha-syn degradation by neurosin between phosphorylated and non-phosphorylated forms of alpha-syn, and between mutant and wild-type alpha-syn. Neurosin cleaved alpha-syn at specific sites. The major cleavage site was localized between Lys80 and Thr81 within the NAC region (E61 to V95), which is important for alpha-syn aggregation, and accordingly may preclude alpha-syn polymerization. Meanwhile, alternative, minor forms of processing also occur. They conserve the NAC region with truncation of the C-terminal region, and accordingly may contribute to alpha-syn polymerization. Phosphorylated alpha-syn was more resistant to degradation by neurosin than non-phosphorylated alpha-syn. The A30P mutant was more resistant to degradation than the wild-type and other alpha-syn mutants. This resistance to neurosin-mediated degradation of phosphorylated alpha-syn and the A30P mutant, which are, respectively, posttranslational and genetic factors related to the development of Parkinson's disease (PD), provides supporting evidence that neurosin is involved in the pathogenesis of PD. (C) 2008 Elsevier Ireland Ltd. All rights reserved.