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基本情報
登録情報 | データベース: EMDB / ID: EMD-2871 | |||||||||
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タイトル | Cryo-EM structure of the Dark apoptosome. | |||||||||
![]() | Reconstruction of Dark apoptosome | |||||||||
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![]() | Cryo-EM / apoptosome / single particle analysis | |||||||||
機能・相同性 | ![]() negative regulation of humoral immune response / positive regulation of glial cell apoptotic process / Formation of apoptosome / salivary gland histolysis / positive regulation of compound eye retinal cell programmed cell death / melanization defense response / sarcosine catabolic process / Activation of caspases through apoptosome-mediated cleavage / Regulation of the apoptosome activity / central nervous system formation ...negative regulation of humoral immune response / positive regulation of glial cell apoptotic process / Formation of apoptosome / salivary gland histolysis / positive regulation of compound eye retinal cell programmed cell death / melanization defense response / sarcosine catabolic process / Activation of caspases through apoptosome-mediated cleavage / Regulation of the apoptosome activity / central nervous system formation / chaeta development / sperm individualization / apoptosome / autophagic cell death / Neutrophil degranulation / S-adenosylmethionine cycle / CARD domain binding / programmed cell death / triglyceride homeostasis / dendrite morphogenesis / response to starvation / cysteine-type endopeptidase activator activity involved in apoptotic process / response to gamma radiation / ADP binding / neuron cellular homeostasis / positive regulation of apoptotic process / ATP binding / identical protein binding 類似検索 - 分子機能 | |||||||||
生物種 | ![]() ![]() | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / ネガティブ染色法 / 解像度: 4.0 Å | |||||||||
![]() | Pang YX / Bai XC / Hao Q / Yan CY / Chen ZQ / Wang JW / Scheres SHW / Shi YG | |||||||||
![]() | ![]() タイトル: Structure of the apoptosome: mechanistic insights into activation of an initiator caspase from Drosophila. 著者: Yuxuan Pang / Xiao-chen Bai / Chuangye Yan / Qi Hao / Zheqin Chen / Jia-Wei Wang / Sjors H W Scheres / Yigong Shi / ![]() ![]() 要旨: Apoptosis is executed by a cascade of caspase activation. The autocatalytic activation of an initiator caspase, exemplified by caspase-9 in mammals or its ortholog, Dronc, in fruit flies, is ...Apoptosis is executed by a cascade of caspase activation. The autocatalytic activation of an initiator caspase, exemplified by caspase-9 in mammals or its ortholog, Dronc, in fruit flies, is facilitated by a multimeric adaptor complex known as the apoptosome. The underlying mechanism by which caspase-9 or Dronc is activated by the apoptosome remains unknown. Here we report the electron cryomicroscopic (cryo-EM) structure of the intact apoptosome from Drosophila melanogaster at 4.0 Å resolution. Analysis of the Drosophila apoptosome, which comprises 16 molecules of the Dark protein (Apaf-1 ortholog), reveals molecular determinants that support the assembly of the 2.5-MDa complex. In the absence of dATP or ATP, Dronc zymogen potently induces formation of the Dark apoptosome, within which Dronc is efficiently activated. At 4.1 Å resolution, the cryo-EM structure of the Dark apoptosome bound to the caspase recruitment domain (CARD) of Dronc (Dronc-CARD) reveals two stacked rings of Dronc-CARD that are sandwiched between two octameric rings of the Dark protein. The specific interactions between Dronc-CARD and both the CARD and the WD40 repeats of a nearby Dark protomer are indispensable for Dronc activation. These findings reveal important mechanistic insights into the activation of initiator caspase by the apoptosome. | |||||||||
履歴 |
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構造の表示
ムービー |
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構造ビューア | EMマップ: ![]() ![]() ![]() |
添付画像 |
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ダウンロードとリンク
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マップデータ | ![]() | 115.7 MB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 9.9 KB 9.9 KB | 表示 表示 | ![]() |
画像 | ![]() | 200.6 KB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
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リンク
EMDBのページ | ![]() ![]() |
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「今月の分子」の関連する項目 |
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マップ
ファイル | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | Reconstruction of Dark apoptosome | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 1.34 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
詳細 | EMDB XML:
CCP4マップ ヘッダ情報:
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-添付データ
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試料の構成要素
-全体 : Dark apoptosome
全体 | 名称: Dark apoptosome |
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要素 |
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-超分子 #1000: Dark apoptosome
超分子 | 名称: Dark apoptosome / タイプ: sample / ID: 1000 / 集合状態: homo 16 mers / Number unique components: 1 |
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分子量 | 実験値: 2.5 MDa / 理論値: 2.5 MDa |
-分子 #1: Dark apoptosome
分子 | 名称: Dark apoptosome / タイプ: protein_or_peptide / ID: 1 / 組換発現: Yes |
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由来(天然) | 生物種: ![]() ![]() 別称: fruit fly / 細胞: Hi-5 insect cells |
分子量 | 実験値: 2.5 MDa / 理論値: 2.5 MDa |
組換発現 | 生物種: ![]() ![]() 組換細胞: Hi-5 insect cells / 組換プラスミド: pFastBac1 |
-実験情報
-構造解析
手法 | ネガティブ染色法, クライオ電子顕微鏡法 |
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![]() | 単粒子再構成法 |
試料の集合状態 | particle |
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試料調製
濃度 | 0.15 mg/mL |
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緩衝液 | pH: 8 / 詳細: 25mM Tris, 150mM NaCl, 5mM dithiothreitol |
染色 | タイプ: NEGATIVE 詳細: Grids were blotted for 2 seconds and flash frozen in liquid ethane using an FEI Vitrobot. |
グリッド | 詳細: Samples were placed on glow-discharged holey carbon grids (Quantifoil CuR2/2), on which a home-made continuous carbon film had previously been deposited. |
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 II / 手法: Blot for 2 seconds before plunging |
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電子顕微鏡法
顕微鏡 | FEI POLARA 300 |
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温度 | 最低: 80 K / 最高: 90 K / 平均: 85 K |
日付 | 2013年11月17日 |
撮影 | カテゴリ: CCD フィルム・検出器のモデル: FEI FALCON II (4k x 4k) デジタル化 - サンプリング間隔: 14 µm / 実像数: 693 / 平均電子線量: 28 e/Å2 詳細: 16 video frames were recorded in 1s by FalconII detector. |
電子線 | 加速電圧: 300 kV / 電子線源: ![]() |
電子光学系 | 倍率(補正後): 104748 / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 2 mm / 最大 デフォーカス(公称値): 6.6 µm / 最小 デフォーカス(公称値): 1.6 µm / 倍率(公称値): 78000 |
試料ステージ | 試料ホルダーモデル: GATAN LIQUID NITROGEN |
実験機器 | ![]() モデル: Tecnai Polara / 画像提供: FEI Company |
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画像解析
CTF補正 | 詳細: Each particle |
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最終 再構成 | 想定した対称性 - 点群: D8 (2回x8回 2面回転対称) 解像度のタイプ: BY AUTHOR / 解像度: 4.0 Å / 解像度の算出法: OTHER / ソフトウェア - 名称: CTFFIND3, RELION 詳細: To correct for beam-induced movements, the 16 video frames for each micrograph were first aligned using whole-image motion correction. Second, particle based beam-induced movement correction ...詳細: To correct for beam-induced movements, the 16 video frames for each micrograph were first aligned using whole-image motion correction. Second, particle based beam-induced movement correction was performed using statistical movie processing in RELION. 使用した粒子像数: 9354 |