Fujiwara Satoru, Araki Katsuya, Matsuo Tatsuhito, Yagi Hisashi, Yamada Takeshi, Shibata Kaoru, Mochizuki Hideki
Quantum Beam Science Center, Japan Atomic Energy Agency, Tokai, Ibaraki, Japan.
Department of Neurology, Osaka University Graduate School of Medicine, Suita, Osaka, Japan.
PLoS One. 2016 Apr 20;11(4):e0151447. doi: 10.1371/journal.pone.0151447. eCollection 2016.
α-synuclein (αSyn) is a protein consisting of 140 amino acid residues and is abundant in the presynaptic nerve terminals in the brain. Although its precise function is unknown, the filamentous aggregates (amyloid fibrils) of αSyn have been shown to be involved in the pathogenesis of Parkinson's disease, which is a progressive neurodegenerative disorder. To understand the pathogenesis mechanism of this disease, the mechanism of the amyloid fibril formation of αSyn must be elucidated. Purified αSyn from bacterial expression is monomeric but intrinsically disordered in solution and forms amyloid fibrils under various conditions. As a first step toward elucidating the mechanism of the fibril formation of αSyn, we investigated dynamical behavior of the purified αSyn in the monomeric state and the fibril state using quasielastic neutron scattering (QENS). We prepared the solution sample of 9.5 mg/ml purified αSyn, and that of 46 mg/ml αSyn in the fibril state, both at pD 7.4 in D2O. The QENS experiments on these samples were performed using the near-backscattering spectrometer, BL02 (DNA), at the Materials and Life Science Facility at the Japan Accelerator Research Complex, Japan. Analysis of the QENS spectra obtained shows that diffusive global motions are observed in the monomeric state but largely suppressed in the fibril state. However, the amplitude of the side chain motion is shown to be larger in the fibril state than in the monomeric state. This implies that significant solvent space exists within the fibrils, which is attributed to the αSyn molecules within the fibrils having a distribution of conformations. The larger amplitude of the side chain motion in the fibril state than in the monomeric state implies that the fibril state is entropically favorable.
α-突触核蛋白（αSyn）是一种由140个氨基酸残基组成的蛋白质，在大脑的突触前神经末梢中含量丰富。尽管其确切功能尚不清楚，但αSyn的丝状聚集体（淀粉样纤维）已被证明与帕金森病的发病机制有关，帕金森病是一种进行性神经退行性疾病。为了理解这种疾病的发病机制，必须阐明αSyn淀粉样纤维形成的机制。从细菌表达中纯化得到的αSyn在溶液中是单体形式但本质上是无序的，并且在各种条件下形成淀粉样纤维。作为阐明αSyn纤维形成机制的第一步，我们使用准弹性中子散射（QENS）研究了纯化的αSyn在单体状态和纤维状态下的动力学行为。我们制备了9.5mg/ml纯化αSyn的溶液样品以及46mg/ml处于纤维状态的αSyn溶液样品，二者均处于重水（D2O）中，pH值为7.4。对这些样品进行的QENS实验是在日本加速器研究综合体材料与生命科学设施的近背散射光谱仪BL02（DNA）上进行的。对所获得的QENS光谱的分析表明，在单体状态下观察到扩散性的整体运动，但在纤维状态下这种运动被大大抑制。然而，侧链运动的幅度在纤维状态下比在单体状态下更大。这意味着在纤维内部存在显著的溶剂空间，这归因于纤维内的αSyn分子具有构象分布。侧链运动在纤维状态下比在单体状态下幅度更大，这意味着纤维状态在熵方面是有利的。
