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基本情報
登録情報 | データベース: PDB / ID: 6roj | |||||||||||||||||||||
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タイトル | Cryo-EM structure of the activated Drs2p-Cdc50p | |||||||||||||||||||||
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![]() | LIPID TRANSPORT / Lipid Flippase / P-type ATPase / PS Transport. | |||||||||||||||||||||
機能・相同性 | ![]() oxaloacetate decarboxylase (Na+ extruding) / decarboxylation-driven active transmembrane transporter activity / Cdc50p-Drs2p complex / actin cortical patch localization / aminophospholipid translocation / Ion transport by P-type ATPases / phosphatidylcholine flippase activity / post-Golgi vesicle-mediated transport / oxaloacetate decarboxylase activity / phosphatidylserine flippase activity ...oxaloacetate decarboxylase (Na+ extruding) / decarboxylation-driven active transmembrane transporter activity / Cdc50p-Drs2p complex / actin cortical patch localization / aminophospholipid translocation / Ion transport by P-type ATPases / phosphatidylcholine flippase activity / post-Golgi vesicle-mediated transport / oxaloacetate decarboxylase activity / phosphatidylserine flippase activity / phospholipid-translocating ATPase complex / ATPase-coupled intramembrane lipid transporter activity / phosphatidylserine floppase activity / phosphatidylethanolamine flippase activity / P-type phospholipid transporter / endocytic recycling / retrograde transport, endosome to Golgi / phosphatidylinositol-4-phosphate binding / phospholipid translocation / sodium ion transport / Neutrophil degranulation / intracellular protein transport / trans-Golgi network / endocytosis / late endosome membrane / endosome membrane / Golgi apparatus / magnesium ion binding / endoplasmic reticulum / ATP hydrolysis activity / ATP binding / plasma membrane / cytosol 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: 2.9 Å | |||||||||||||||||||||
![]() | Timcenko, M. / Lyons, J.A. / Januliene, D. / Ulstrup, J.J. / Dieudonne, T. / Montigny, C. / Ash, M.R. / Karlsen, J.L. / Boesen, T. / Kuhlbrandt, W. ...Timcenko, M. / Lyons, J.A. / Januliene, D. / Ulstrup, J.J. / Dieudonne, T. / Montigny, C. / Ash, M.R. / Karlsen, J.L. / Boesen, T. / Kuhlbrandt, W. / Lenoir, G. / Moeller, A. / Nissen, P. | |||||||||||||||||||||
資金援助 | ![]() ![]()
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![]() | ![]() タイトル: Structure and autoregulation of a P4-ATPase lipid flippase. 著者: Milena Timcenko / Joseph A Lyons / Dovile Januliene / Jakob J Ulstrup / Thibaud Dieudonné / Cédric Montigny / Miriam-Rose Ash / Jesper Lykkegaard Karlsen / Thomas Boesen / Werner ...著者: Milena Timcenko / Joseph A Lyons / Dovile Januliene / Jakob J Ulstrup / Thibaud Dieudonné / Cédric Montigny / Miriam-Rose Ash / Jesper Lykkegaard Karlsen / Thomas Boesen / Werner Kühlbrandt / Guillaume Lenoir / Arne Moeller / Poul Nissen / ![]() ![]() ![]() 要旨: Type 4 P-type ATPases (P4-ATPases) are lipid flippases that drive the active transport of phospholipids from exoplasmic or luminal leaflets to cytosolic leaflets of eukaryotic membranes. The ...Type 4 P-type ATPases (P4-ATPases) are lipid flippases that drive the active transport of phospholipids from exoplasmic or luminal leaflets to cytosolic leaflets of eukaryotic membranes. The molecular architecture of P4-ATPases and the mechanism through which they recognize and transport lipids have remained unknown. Here we describe the cryo-electron microscopy structure of the P4-ATPase Drs2p-Cdc50p, a Saccharomyces cerevisiae lipid flippase that is specific to phosphatidylserine and phosphatidylethanolamine. Drs2p-Cdc50p is autoinhibited by the C-terminal tail of Drs2p, and activated by the lipid phosphatidylinositol-4-phosphate (PtdIns4P or PI4P). We present three structures that represent the complex in an autoinhibited, an intermediate and a fully activated state. The analysis highlights specific features of P4-ATPases and reveals sites of autoinhibition and PI4P-dependent activation. We also observe a putative lipid translocation pathway in this flippase that involves a conserved PISL motif in transmembrane segment 4 and polar residues of transmembrane segments 2 and 5, in particular Lys1018, in the centre of the lipid bilayer. | |||||||||||||||||||||
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構造の表示
ムービー |
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構造ビューア | 分子: ![]() ![]() |
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PDBx/mmCIF形式 | ![]() | 269.8 KB | 表示 | ![]() |
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PDB形式 | ![]() | 213.9 KB | 表示 | ![]() |
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その他 | ![]() |
-検証レポート
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
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-関連構造データ
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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
-タンパク質 , 2種, 2分子 AC
#1: タンパク質 | 分子量: 164240.359 Da / 分子数: 1 / 由来タイプ: 組換発現 詳細: Residues 1-104 removed after proteolysis with thrombin Residues 1252-1364 removed after proteolysis with thrombin 1248-1253 - added an addition thrombin cleavage site D560 described by ...詳細: Residues 1-104 removed after proteolysis with thrombin Residues 1252-1364 removed after proteolysis with thrombin 1248-1253 - added an addition thrombin cleavage site D560 described by Aspartate beryllium trifluoride (BFD) engineered C-terminal GGGG-LVPRGS-BAD-ta,Residues 1-104 removed after proteolysis with thrombin Residues 1252-1364 removed after proteolysis with thrombin 1248-1253 - added an addition thrombin cleavage site D560 described by Aspartate beryllium trifluoride (BFD) engineered C-terminal GGGG-LVPRGS-BAD-ta 由来: (組換発現) ![]() ![]() ![]() 遺伝子: DRS2, YAL026C, FUN38, oadA / プラスミド: pYedp60 / 発現宿主: ![]() ![]() 参照: UniProt: P39524, UniProt: P13187, P-type phospholipid transporter, oxaloacetate decarboxylase (Na+ extruding) |
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#2: タンパク質 | 分子量: 47371.797 Da / 分子数: 1 / 由来タイプ: 組換発現 / 詳細: engineered C-terminal GGGG-LVPRGS-GG-10xHistag 由来: (組換発現) ![]() ![]() 遺伝子: CDC50, YCR094W, YCR94W / プラスミド: pYedp60 / 発現宿主: ![]() ![]() |
-糖 , 2種, 3分子
#3: 多糖 | #4: 多糖 | 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose | |
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-非ポリマー , 3種, 4分子 ![](data/chem/img/MG.gif)
![](data/chem/img/2Y5.gif)
![](data/chem/img/HOH.gif)
![](data/chem/img/2Y5.gif)
![](data/chem/img/HOH.gif)
#5: 化合物 | ChemComp-MG / |
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#6: 化合物 | ChemComp-2Y5 / ( |
#7: 水 | ChemComp-HOH / |
-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 | 名称: Binary complex of Drs2p-Cdc50p with the regulatory lipid PI4P タイプ: COMPLEX 詳細: C-terminus removed by proteolytic cleavage of an engineered site. Entity ID: #1-#2 / 由来: RECOMBINANT |
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分子量 | 実験値: NO |
由来(天然) | 生物種: ![]() ![]() |
由来(組換発現) | 生物種: ![]() ![]() |
緩衝液 | pH: 7 |
試料 | 濃度: 0.6 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES / 詳細: 1mM beryllium fluoride 0.1 mg/mL Brain PI4P in DDM |
試料支持 | 詳細: 15mA / グリッドの材料: COPPER / グリッドのサイズ: 400 divisions/in. / グリッドのタイプ: C-flat-1.2/1.3 |
急速凍結 | 装置: FEI VITROBOT MARK IV / 凍結剤: ETHANE / 湿度: 100 % / 凍結前の試料温度: 278 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 |
撮影 | 平均露光時間: 8 sec. / 電子線照射量: 56 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) |
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解析
ソフトウェア | 名称: PHENIX / バージョン: 1.14_3260: / 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: 1047615 | |||||||||||||||||||||
対称性 | 点対称性: C1 (非対称) | |||||||||||||||||||||
3次元再構成 | 解像度: 2.9 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 418512 / 対称性のタイプ: POINT |