+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-2846 | |||||||||
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タイトル | Three-dimensional structure of the autophagic phosphatidylinositol 3-kinase complex. | |||||||||
マップデータ | Three-dimensional structure of the autophagic phosphatidylinositol 3-kinase complex, PI3KC3-C1. | |||||||||
試料 |
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キーワード | kinase / lipid kinase / protein kinase / autophagy / phosphatidylinositol 3-kinase / PI3K / phosphatidylinositol 3-phosphate / PI(3)P | |||||||||
機能・相同性 | 機能・相同性情報 extrinsic component of omegasome membrane / phosphatidylinositol 3-kinase inhibitor activity / regulation of triglyceride metabolic process / extrinsic component of phagophore assembly site membrane / nucleus-vacuole junction / cellular response to aluminum ion / Toll Like Receptor 9 (TLR9) Cascade / protein lipidation / Synthesis of PIPs at the late endosome membrane / phosphatidylinositol 3-kinase complex, class III ...extrinsic component of omegasome membrane / phosphatidylinositol 3-kinase inhibitor activity / regulation of triglyceride metabolic process / extrinsic component of phagophore assembly site membrane / nucleus-vacuole junction / cellular response to aluminum ion / Toll Like Receptor 9 (TLR9) Cascade / protein lipidation / Synthesis of PIPs at the late endosome membrane / phosphatidylinositol 3-kinase complex, class III / Synthesis of PIPs at the early endosome membrane / cellular response to oxygen-glucose deprivation / phosphatidylinositol 3-kinase complex, class III, type II / positive regulation of stress granule assembly / phosphatidylinositol 3-kinase complex, class III, type I / mitochondria-associated endoplasmic reticulum membrane contact site / autophagy of peroxisome / response to mitochondrial depolarisation / positive regulation of attachment of mitotic spindle microtubules to kinetochore / cytoplasmic side of mitochondrial outer membrane / negative regulation of lysosome organization / positive regulation by host of viral genome replication / Synthesis of PIPs at the Golgi membrane / engulfment of apoptotic cell / positive regulation of autophagosome assembly / receptor catabolic process / negative regulation of autophagosome assembly / regulation of protein complex stability / phosphatidylinositol kinase activity / phosphatidylinositol 3-kinase regulator activity / protein localization to phagophore assembly site / suppression by virus of host autophagy / protein targeting to vacuole / protein targeting to lysosome / phagophore assembly site membrane / late endosome to vacuole transport / early endosome to late endosome transport / SMAD protein signal transduction / negative regulation of programmed cell death / Translation of Replicase and Assembly of the Replication Transcription Complex / phagophore assembly site / cellular response to nitrogen starvation / response to iron(II) ion / phosphatidylinositol 3-kinase / phosphatidylinositol-3-phosphate biosynthetic process / autophagosome membrane docking / cytoplasmic pattern recognition receptor signaling pathway / mitotic metaphase chromosome alignment / post-transcriptional regulation of gene expression / 1-phosphatidylinositol-3-kinase activity / endosome to lysosome transport / lysosome organization / Macroautophagy / RSV-host interactions / positive regulation of cardiac muscle hypertrophy / autolysosome / p38MAPK cascade / phosphatidylinositol-mediated signaling / axoneme / phosphatidylinositol phosphate biosynthetic process / autophagosome membrane / autophagosome maturation / autophagosome assembly / PI3K Cascade / mitophagy / RHO GTPases Activate NADPH Oxidases / response to vitamin E / negative regulation of reactive oxygen species metabolic process / neuron development / amyloid-beta metabolic process / regulation of macroautophagy / cellular defense response / phosphatidylinositol 3-kinase binding / cellular response to glucose starvation / positive regulation of autophagy / phagocytic vesicle / positive regulation of intrinsic apoptotic signaling pathway / protein-membrane adaptor activity / JNK cascade / cellular response to epidermal growth factor stimulus / cellular response to copper ion / autophagosome / phosphatidylinositol 3-kinase/protein kinase B signal transduction / cellular response to amino acid starvation / cellular response to starvation / regulation of cytokinesis / negative regulation of protein phosphorylation / regulation of autophagy / macroautophagy / Antigen Presentation: Folding, assembly and peptide loading of class I MHC / response to lead ion / regulation of protein phosphorylation / trans-Golgi network / ISG15 antiviral mechanism / autophagy / cellular response to hydrogen peroxide / endocytosis / phagocytic vesicle membrane / microtubule cytoskeleton / late endosome 類似検索 - 分子機能 | |||||||||
生物種 | Homo sapiens (ヒト) | |||||||||
手法 | 単粒子再構成法 / ネガティブ染色法 / 解像度: 27.5 Å | |||||||||
データ登録者 | Baskaran S / Carlson L-A / Stjepanovic G / Young LN / Kim DJ / Grob P / Stanley RE / Nogales E / Hurley JH | |||||||||
引用 | ジャーナル: Elife / 年: 2014 タイトル: Architecture and dynamics of the autophagic phosphatidylinositol 3-kinase complex. 著者: Sulochanadevi Baskaran / Lars-Anders Carlson / Goran Stjepanovic / Lindsey N Young / Do Jin Kim / Patricia Grob / Robin E Stanley / Eva Nogales / James H Hurley / 要旨: The class III phosphatidylinositol 3-kinase complex I (PI3KC3-C1) that functions in early autophagy consists of the lipid kinase VPS34, the scaffolding protein VPS15, the tumor suppressor BECN1, and ...The class III phosphatidylinositol 3-kinase complex I (PI3KC3-C1) that functions in early autophagy consists of the lipid kinase VPS34, the scaffolding protein VPS15, the tumor suppressor BECN1, and the autophagy-specific subunit ATG14. The structure of the ATG14-containing PI3KC3-C1 was determined by single-particle EM, revealing a V-shaped architecture. All of the ordered domains of VPS34, VPS15, and BECN1 were mapped by MBP tagging. The dynamics of the complex were defined using hydrogen-deuterium exchange, revealing a novel 20-residue ordered region C-terminal to the VPS34 C2 domain. VPS15 organizes the complex and serves as a bridge between VPS34 and the ATG14:BECN1 subcomplex. Dynamic transitions occur in which the lipid kinase domain is ejected from the complex and VPS15 pivots at the base of the V. The N-terminus of BECN1, the target for signaling inputs, resides near the pivot point. These observations provide a framework for understanding the allosteric regulation of lipid kinase activity. | |||||||||
履歴 |
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-構造の表示
ムービー |
ムービービューア |
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構造ビューア | EMマップ: SurfViewMolmilJmol/JSmol |
添付画像 |
-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_2846.map.gz | 3.3 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-2846-v30.xml emd-2846.xml | 15.3 KB 15.3 KB | 表示 表示 | EMDBヘッダ |
画像 | emd_2846.tif | 86.5 KB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-2846 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2846 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_2846_validation.pdf.gz | 199 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_2846_full_validation.pdf.gz | 198.1 KB | 表示 | |
XML形式データ | emd_2846_validation.xml.gz | 5.4 KB | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2846 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2846 | HTTPS FTP |
-関連構造データ
-リンク
EMDBのページ | EMDB (EBI/PDBe) / EMDataResource |
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「今月の分子」の関連する項目 |
-マップ
ファイル | ダウンロード / ファイル: emd_2846.map.gz / 形式: CCP4 / 大きさ: 7.8 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | Three-dimensional structure of the autophagic phosphatidylinositol 3-kinase complex, PI3KC3-C1. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
投影像・断面図 | 画像のコントロール
画像は Spider により作成 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 3.01 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
-試料の構成要素
-全体 : Autophagic phosphatidylinositol 3-kinase complex, PI3KC3-C1.
全体 | 名称: Autophagic phosphatidylinositol 3-kinase complex, PI3KC3-C1. |
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要素 |
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-超分子 #1000: Autophagic phosphatidylinositol 3-kinase complex, PI3KC3-C1.
超分子 | 名称: Autophagic phosphatidylinositol 3-kinase complex, PI3KC3-C1. タイプ: sample / ID: 1000 / 集合状態: heterotetramer / Number unique components: 4 |
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分子量 | 理論値: 361.8 KDa |
-分子 #1: phosphatidylinositol 3-kinase, catalytic subunit type 3
分子 | 名称: phosphatidylinositol 3-kinase, catalytic subunit type 3 タイプ: protein_or_peptide / ID: 1 / Name.synonym: VPS34, PI3KC3 / コピー数: 1 / 集合状態: one component of heterotetramer / 組換発現: Yes |
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由来(天然) | 生物種: Homo sapiens (ヒト) / 別称: Human / Organelle: autophagosome / 細胞中の位置: autophagosome membrane |
分子量 | 理論値: 102 KDa |
組換発現 | 生物種: Homo sapiens (ヒト) / 組換細胞: HEK293 / 組換プラスミド: pCAG |
配列 | UniProtKB: Phosphatidylinositol 3-kinase catalytic subunit type 3 GO: 1-phosphatidylinositol-3-kinase activity, phosphatidylinositol 3-kinase complex, class III, type I, autophagy, autophagosome assembly |
-分子 #2: phosphoinositide-3-kinase, regulatory subunit 4
分子 | 名称: phosphoinositide-3-kinase, regulatory subunit 4 / タイプ: protein_or_peptide / ID: 2 / Name.synonym: VPS15, p150, PIK3R4 / コピー数: 1 / 集合状態: one component of heterotetramer / 組換発現: Yes |
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由来(天然) | 生物種: Homo sapiens (ヒト) / 別称: Human / Organelle: autophagosome / 細胞中の位置: autophagosome membrane |
分子量 | 理論値: 153 KDa |
組換発現 | 生物種: Homo sapiens (ヒト) / 組換細胞: HEK293 / 組換プラスミド: pCAG |
配列 | UniProtKB: Phosphoinositide 3-kinase regulatory subunit 4 GO: phosphatidylinositol 3-kinase complex, class III, type I, autophagy, autophagosome assembly |
-分子 #3: beclin1
分子 | 名称: beclin1 / タイプ: protein_or_peptide / ID: 3 / Name.synonym: BECN1 / コピー数: 1 / 集合状態: one component of heterotetramer / 組換発現: Yes |
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由来(天然) | 生物種: Homo sapiens (ヒト) / 別称: Human / Organelle: autophagosome / 細胞中の位置: autophagosome membrane |
分子量 | 理論値: 52 KDa |
組換発現 | 生物種: Homo sapiens (ヒト) / 組換細胞: HEK293 / 組換プラスミド: pCAG |
配列 | UniProtKB: Beclin-1 GO: phosphatidylinositol 3-kinase complex, class III, type I, autophagy, autophagosome assembly |
-分子 #4: autophagy related 14
分子 | 名称: autophagy related 14 / タイプ: protein_or_peptide / ID: 4 / Name.synonym: ATG14, ATG14L, BARKOR / コピー数: 1 / 集合状態: one component of heterotetramer / 組換発現: Yes |
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由来(天然) | 生物種: Homo sapiens (ヒト) / 別称: Human / Organelle: autophagosome / 細胞中の位置: autophagosome membrane |
分子量 | 理論値: 55 KDa |
組換発現 | 生物種: Homo sapiens (ヒト) / 組換細胞: HEK293 / 組換プラスミド: pCAG |
配列 | UniProtKB: Beclin 1-associated autophagy-related key regulator GO: phosphatidylinositol 3-kinase complex, class III, type I, autophagy, autophagosome assembly |
-実験情報
-構造解析
手法 | ネガティブ染色法 |
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解析 | 単粒子再構成法 |
試料の集合状態 | particle |
-試料調製
濃度 | 0.01 mg/mL |
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緩衝液 | pH: 8 詳細: 20 mM Tris-HCl, 200 mM NaCl, 2 mM MgCl2, 1 mM TCEP, 3% trehalose |
染色 | タイプ: NEGATIVE 詳細: Protein was incubated on grids for 30s, followed by sequential 10 s incubations on four 50 ul drops of 1% uranyl formate. The stained grids were blotted to near dryness with a filter paper and air-dried. |
グリッド | 詳細: Continuous carbon, plasma cleaned for 10 s in a Gatan Solarus at 10% O2. |
凍結 | 凍結剤: NONE / 装置: OTHER |
-電子顕微鏡法
顕微鏡 | FEI TECNAI F20 |
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温度 | 最低: 292 K / 最高: 294 K / 平均: 293 K |
アライメント法 | Legacy - 非点収差: Objective lens astigmatism was corrected at 80,000 times magnification. |
日付 | 2014年5月28日 |
撮影 | カテゴリ: CCD フィルム・検出器のモデル: GATAN ULTRASCAN 4000 (4k x 4k) デジタル化 - サンプリング間隔: 15 µm / 実像数: 1614 / 平均電子線量: 25 e/Å2 / ビット/ピクセル: 32 |
Tilt angle min | 0 |
電子線 | 加速電圧: 120 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | 倍率(補正後): 100333 / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 2.2 mm / 最大 デフォーカス(公称値): 3.0 µm / 最小 デフォーカス(公称値): 0.5 µm / 倍率(公称値): 80000 |
試料ステージ | 試料ホルダーモデル: SIDE ENTRY, EUCENTRIC / Tilt angle max: 45 |
実験機器 | モデル: Tecnai F20 / 画像提供: FEI Company |
-画像解析
詳細 | The final three-dimensional structure was calculated in RELION using a single 3D class. Starting model for this calculation was a random conical tilt model low-pass filtered to 80A. |
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CTF補正 | 詳細: tilt-dependent phase flip |
最終 再構成 | 想定した対称性 - 点群: C1 (非対称) / アルゴリズム: OTHER / 解像度のタイプ: BY AUTHOR / 解像度: 27.5 Å / 解像度の算出法: OTHER ソフトウェア - 名称: Spider, EMAN, EMAN2, IMAGIC, RELION 詳細: Final map calculated in RELION using a single 3D class. Data acquired at 0, 30, and 45 degrees were used to refine an initial RCT model low-pass filtered to 80A. Gold standard FSC calculated using RELION. 使用した粒子像数: 38745 |