ジャーナル: Nature / 年: 2015 タイトル: An atomic structure of human γ-secretase. 著者: Xiao-Chen Bai / Chuangye Yan / Guanghui Yang / Peilong Lu / Dan Ma / Linfeng Sun / Rui Zhou / Sjors H W Scheres / Yigong Shi / 要旨: Dysfunction of the intramembrane protease γ-secretase is thought to cause Alzheimer's disease, with most mutations derived from Alzheimer's disease mapping to the catalytic subunit presenilin 1 (PS1) ...Dysfunction of the intramembrane protease γ-secretase is thought to cause Alzheimer's disease, with most mutations derived from Alzheimer's disease mapping to the catalytic subunit presenilin 1 (PS1). Here we report an atomic structure of human γ-secretase at 3.4 Å resolution, determined by single-particle cryo-electron microscopy. Mutations derived from Alzheimer's disease affect residues at two hotspots in PS1, each located at the centre of a distinct four transmembrane segment (TM) bundle. TM2 and, to a lesser extent, TM6 exhibit considerable flexibility, yielding a plastic active site and adaptable surrounding elements. The active site of PS1 is accessible from the convex side of the TM horseshoe, suggesting considerable conformational changes in nicastrin extracellular domain after substrate recruitment. Component protein APH-1 serves as a scaffold, anchoring the lone transmembrane helix from nicastrin and supporting the flexible conformation of PS1. Ordered phospholipids stabilize the complex inside the membrane. Our structure serves as a molecular basis for mechanistic understanding of γ-secretase function.
EMPIAR-10194 (タイトル: An atomic structure of human gamma-secretase / Data size: 11.7 TB Data #1: Unaligned multi-frame micrographs of human gamma-secretase [micrographs - multiframe])
pH: 7.4 / 詳細: 25 mM HEPES, pH 7.4, 150 mM NaCl and amphipol A8-35
染色
タイプ: NEGATIVE / 詳細: cryo-EM
グリッド
詳細: 300 mesh Au 1.2/1.3 Quantifoil grid, glow discharged for 1 minute
凍結
凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 85 K / 装置: FEI VITROBOT MARK IV / 手法: Blot for 4 seconds before plunging
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電子顕微鏡法
顕微鏡
FEI TITAN KRIOS
温度
最低: 80 K / 最高: 90 K / 平均: 85 K
特殊光学系
エネルギーフィルター - 名称: Gatan Quantum エネルギーフィルター - エネルギー下限: 0.0 eV エネルギーフィルター - エネルギー上限: 20.0 eV
日付
2014年10月2日
撮影
カテゴリ: CCD / フィルム・検出器のモデル: GATAN K2 (4k x 4k) / デジタル化 - サンプリング間隔: 5 µm / 実像数: 2925 / 平均電子線量: 38 e/Å2 詳細: 20 frames were recorded by the direct electron detector
Use a newly developed statistical movie processing and particle polishing approach to compensate for beam-induced movement and reduce the effect of radiation-damage.
CTF補正
詳細: Each particle
最終 再構成
想定した対称性 - 点群: C1 (非対称) / 解像度のタイプ: BY AUTHOR / 解像度: 3.4 Å / 解像度の算出法: OTHER / ソフトウェア - 名称: CTFFIND3, RELION / 使用した粒子像数: 159549