+Open data
-Basic information
Entry | Database: PDB / ID: 6roi | |||||||||||||||||||||
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Title | Cryo-EM structure of the partially activated Drs2p-Cdc50p | |||||||||||||||||||||
Components |
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Keywords | LIPID TRANSPORT / Lipid Flippase / P-type ATPase / PS Transport. | |||||||||||||||||||||
Function / homology | Function and homology information Cdc50p-Drs2p complex / actin cortical patch localization / Ion transport by P-type ATPases / phosphatidylcholine flippase activity / aminophospholipid translocation / post-Golgi vesicle-mediated transport / phosphatidylserine flippase activity / ATPase-coupled intramembrane lipid transporter activity / phosphatidylserine floppase activity / phospholipid-translocating ATPase complex ...Cdc50p-Drs2p complex / actin cortical patch localization / Ion transport by P-type ATPases / phosphatidylcholine flippase activity / aminophospholipid translocation / post-Golgi vesicle-mediated transport / phosphatidylserine flippase activity / ATPase-coupled intramembrane lipid transporter activity / phosphatidylserine floppase activity / phospholipid-translocating ATPase complex / phosphatidylethanolamine flippase activity / P-type phospholipid transporter / endocytic recycling / retrograde transport, endosome to Golgi / phosphatidylinositol-4-phosphate binding / phospholipid translocation / 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 Similarity search - Function | |||||||||||||||||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | |||||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å | |||||||||||||||||||||
Authors | 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. | |||||||||||||||||||||
Funding support | Denmark, France, 6items
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Citation | Journal: Nature / Year: 2019 Title: Structure and autoregulation of a P4-ATPase lipid flippase. Authors: Milena Timcenko / Joseph A Lyons / Dovile Januliene / Jakob J Ulstrup / Thibaud Dieudonné / Cédric Montigny / Miriam-Rose Ash / Jesper Lykkegaard Karlsen / Thomas Boesen / Werner ...Authors: 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 / Abstract: 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. | |||||||||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6roi.cif.gz | 276.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6roi.ent.gz | 220 KB | Display | PDB format |
PDBx/mmJSON format | 6roi.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6roi_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 6roi_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 6roi_validation.xml.gz | 54.4 KB | Display | |
Data in CIF | 6roi_validation.cif.gz | 80.6 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ro/6roi ftp://data.pdbj.org/pub/pdb/validation_reports/ro/6roi | HTTPS FTP |
-Related structure data
Related structure data | 4973MC 4972C 4974C 6rohC 6rojC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein , 2 types, 2 molecules AC
#1: Protein | Mass: 163730.766 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: D560 described by Aspartate beryllium trifluoride (BFD) engineered C-terminal GGGG-LVPRGS-BAD-tag Residues 1-104 are removed by proteolysis with thrombin Residues 1364-1460 are removed after ...Details: D560 described by Aspartate beryllium trifluoride (BFD) engineered C-terminal GGGG-LVPRGS-BAD-tag Residues 1-104 are removed by proteolysis with thrombin Residues 1364-1460 are removed after cleaving of the purification tag by thrombin Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / Gene: DRS2, YAL026C, FUN38 / Production host: Saccharomyces cerevisiae S288C (yeast) References: UniProt: P39524, P-type phospholipid transporter |
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#2: Protein | Mass: 47371.797 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: engineered C-terminal GGGG-LVPRGS-GG-10xHistag Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / Gene: CDC50, YCR094W, YCR94W / Production host: Saccharomyces cerevisiae S288C (yeast) / References: UniProt: P25656 |
-Sugars , 3 types, 3 molecules
#3: Polysaccharide | 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source |
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#4: Polysaccharide | beta-D-mannopyranose-(1-3)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4) ...beta-D-mannopyranose-(1-3)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source |
#7: Sugar | ChemComp-NAG / |
-Non-polymers , 3 types, 4 molecules
#5: Chemical | ChemComp-MG / |
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#6: Chemical | ChemComp-2Y5 / ( |
#8: Water | ChemComp-HOH / |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Binary complex of Drs2p-Cdc50p with the regulatory lipid PI4P Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Saccharomyces cerevisiae S288C (yeast) |
Source (recombinant) | Organism: Saccharomyces cerevisiae S288C (yeast) / Plasmid: pYedp60 |
Buffer solution | pH: 7 |
Specimen | Conc.: 0.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: 1mM beryllium fluoride 0.1 mg/mL Brain PI4P in DDM |
Specimen support | Details: 15 mA / Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: C-flat-1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 278 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 60 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) |
-Processing
Software | Name: PHENIX / Version: 1.14_3260: / Classification: refinement | |||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||
Particle selection | Num. of particles selected: 578440 | |||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||
3D reconstruction | Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 78981 / Num. of class averages: 1 / Symmetry type: POINT |