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基本情報
登録情報 | データベース: PDB / ID: 8cr1 | |||||||||
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タイトル | Homo sapiens Get1/Get2 heterotetramer in complex with a Get3 dimer | |||||||||
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![]() | MEMBRANE PROTEIN / membrane protein insertion / GET pathway / tail anchored membrane protein | |||||||||
機能・相同性 | ![]() arsenite transmembrane transporter activity / otic vesicle development / membrane insertase activity / GET complex / tail-anchored membrane protein insertion into ER membrane / receptor recycling / protein insertion into ER membrane / post-translational protein targeting to endoplasmic reticulum membrane / Insertion of tail-anchored proteins into the endoplasmic reticulum membrane / B cell homeostasis ...arsenite transmembrane transporter activity / otic vesicle development / membrane insertase activity / GET complex / tail-anchored membrane protein insertion into ER membrane / receptor recycling / protein insertion into ER membrane / post-translational protein targeting to endoplasmic reticulum membrane / Insertion of tail-anchored proteins into the endoplasmic reticulum membrane / B cell homeostasis / vesicle-mediated transport / protein-membrane adaptor activity / negative regulation of proteasomal ubiquitin-dependent protein catabolic process / negative regulation of protein ubiquitination / establishment of localization in cell / sensory perception of sound / synapse organization / defense response / 加水分解酵素; 酸無水物に作用; 酸無水物に作用・細胞または細胞小器官の運動に関与 / epidermal growth factor receptor signaling pathway / protein stabilization / ubiquitin protein ligase binding / endoplasmic reticulum membrane / nucleolus / endoplasmic reticulum / signal transduction / ATP hydrolysis activity / extracellular exosome / nucleoplasm / ATP binding / metal ion binding / nucleus / membrane / cytoplasm 類似検索 - 分子機能 | |||||||||
生物種 | ![]() | |||||||||
手法 | 電子顕微鏡法 / 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 3.2 Å | |||||||||
![]() | McDowell, M.A. / Heimes, M. / Wild, K. / Sinning, I. | |||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: The GET insertase exhibits conformational plasticity and induces membrane thinning. 著者: Melanie A McDowell / Michael Heimes / Giray Enkavi / Ákos Farkas / Daniel Saar / Klemens Wild / Blanche Schwappach / Ilpo Vattulainen / Irmgard Sinning / ![]() ![]() 要旨: The eukaryotic guided entry of tail-anchored proteins (GET) pathway mediates the biogenesis of tail-anchored (TA) membrane proteins at the endoplasmic reticulum. In the cytosol, the Get3 chaperone ...The eukaryotic guided entry of tail-anchored proteins (GET) pathway mediates the biogenesis of tail-anchored (TA) membrane proteins at the endoplasmic reticulum. In the cytosol, the Get3 chaperone captures the TA protein substrate and delivers it to the Get1/Get2 membrane protein complex (GET insertase), which then inserts the substrate via a membrane-embedded hydrophilic groove. Here, we present structures, atomistic simulations and functional data of human and Chaetomium thermophilum Get1/Get2/Get3. The core fold of the GET insertase is conserved throughout eukaryotes, whilst thinning of the lipid bilayer occurs in the vicinity of the hydrophilic groove to presumably lower the energetic barrier of membrane insertion. We show that the gating interaction between Get2 helix α3' and Get3 drives conformational changes in both Get3 and the Get1/Get2 membrane heterotetramer. Thus, we provide a framework to understand the conformational plasticity of the GET insertase and how it remodels its membrane environment to promote substrate insertion. #1: ![]() タイトル: Structural Basis of Tail-Anchored Membrane Protein Biogenesis by the GET Insertase Complex. 著者: Melanie A McDowell / Michael Heimes / Francesco Fiorentino / Shahid Mehmood / Ákos Farkas / Javier Coy-Vergara / Di Wu / Jani Reddy Bolla / Volker Schmid / Roger Heinze / Klemens Wild / Dirk ...著者: Melanie A McDowell / Michael Heimes / Francesco Fiorentino / Shahid Mehmood / Ákos Farkas / Javier Coy-Vergara / Di Wu / Jani Reddy Bolla / Volker Schmid / Roger Heinze / Klemens Wild / Dirk Flemming / Stefan Pfeffer / Blanche Schwappach / Carol V Robinson / Irmgard Sinning / ![]() ![]() 要旨: Membrane protein biogenesis faces the challenge of chaperoning hydrophobic transmembrane helices for faithful membrane insertion. The guided entry of tail-anchored proteins (GET) pathway targets and ...Membrane protein biogenesis faces the challenge of chaperoning hydrophobic transmembrane helices for faithful membrane insertion. The guided entry of tail-anchored proteins (GET) pathway targets and inserts tail-anchored (TA) proteins into the endoplasmic reticulum (ER) membrane with an insertase (yeast Get1/Get2 or mammalian WRB/CAML) that captures the TA from a cytoplasmic chaperone (Get3 or TRC40, respectively). Here, we present cryo-electron microscopy reconstructions, native mass spectrometry, and structure-based mutagenesis of human WRB/CAML/TRC40 and yeast Get1/Get2/Get3 complexes. Get3 binding to the membrane insertase supports heterotetramer formation, and phosphatidylinositol binding at the heterotetramer interface stabilizes the insertase for efficient TA insertion in vivo. We identify a Get2/CAML cytoplasmic helix that forms a "gating" interaction with Get3/TRC40 important for TA insertion. Structural homology with YidC and the ER membrane protein complex (EMC) implicates an evolutionarily conserved insertion mechanism for divergent substrates utilizing a hydrophilic groove. Thus, we provide a detailed structural and mechanistic framework to understand TA membrane insertion. | |||||||||
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構造の表示
構造ビューア | 分子: ![]() ![]() |
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ダウンロードとリンク
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PDBx/mmCIF形式 | ![]() | 214.3 KB | 表示 | ![]() |
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PDB形式 | ![]() | 168.9 KB | 表示 | ![]() |
PDBx/mmJSON形式 | ![]() | ツリー表示 | ![]() | |
その他 | ![]() |
-検証レポート
文書・要旨 | ![]() | 1.1 MB | 表示 | ![]() |
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文書・詳細版 | ![]() | 1.1 MB | 表示 | |
XML形式データ | ![]() | 49.8 KB | 表示 | |
CIF形式データ | ![]() | 72.5 KB | 表示 | |
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
関連構造データ | ![]() 16801MC ![]() 8cqzC ![]() 8cr2C ![]() 8oduC ![]() 8odvC M: このデータのモデリングに利用したマップデータ C: 同じ文献を引用 ( |
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類似構造データ | 類似検索 - 機能・相同性 ![]() |
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リンク
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集合体
登録構造単位 | ![]()
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要素
#1: タンパク質 | 分子量: 40146.070 Da / 分子数: 2 / 由来タイプ: 組換発現 / 由来: (組換発現) ![]() ![]() ![]() 参照: UniProt: O43681, 加水分解酵素; 酸無水物に作用 #2: タンパク質 | 分子量: 35083.309 Da / 分子数: 2 / Mutation: Truncation of 185 N-terminal residues. / 由来タイプ: 組換発現 / 由来: (組換発現) ![]() 発現宿主: ![]() ![]() 参照: UniProt: P49069, UniProt: O00258 #3: 化合物 | ChemComp-ZN / | 研究の焦点であるリガンドがあるか | N | Has protein modification | Y | |
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-実験情報
-実験
実験 | 手法: 電子顕微鏡法 |
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EM実験 | 試料の集合状態: PARTICLE / 3次元再構成法: 単粒子再構成法 |
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試料調製
構成要素 |
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分子量 | 値: 0.150 MDa / 実験値: NO | ||||||||||||||||||||||||||||
由来(天然) |
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由来(組換発現) |
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緩衝液 | pH: 7.5 | ||||||||||||||||||||||||||||
緩衝液成分 |
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試料 | 濃度: 1.8 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES / 詳細: Complex stabilised in PMAL-C8 amphipol | ||||||||||||||||||||||||||||
試料支持 | グリッドの材料: COPPER/RHODIUM / グリッドのサイズ: 300 divisions/in. / グリッドのタイプ: Quantifoil R2/1 | ||||||||||||||||||||||||||||
急速凍結 | 装置: FEI VITROBOT MARK IV / 凍結剤: ETHANE / 湿度: 95 % / 凍結前の試料温度: 279 K |
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電子顕微鏡撮影
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TITAN KRIOS |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 最大 デフォーカス(公称値): 2000 nm / 最小 デフォーカス(公称値): 800 nm / Cs: 2.7 mm / C2レンズ絞り径: 50 µm |
試料ホルダ | 凍結剤: NITROGEN |
撮影 | 平均露光時間: 12 sec. / 電子線照射量: 45.6 e/Å2 / 検出モード: COUNTING フィルム・検出器のモデル: GATAN K2 QUANTUM (4k x 4k) 撮影したグリッド数: 1 / 実像数: 9470 |
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解析
ソフトウェア | 名称: PHENIX / バージョン: 1.19_4092: / 分類: 精密化 | ||||||||||||||||||||||||||||||||||||
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EMソフトウェア |
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CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
粒子像の選択 | 選択した粒子像数: 1561837 | ||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C2 (2回回転対称) | ||||||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 3.2 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 189844 / 対称性のタイプ: POINT | ||||||||||||||||||||||||||||||||||||
原子モデル構築 | プロトコル: RIGID BODY FIT / 空間: REAL | ||||||||||||||||||||||||||||||||||||
拘束条件 |
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