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Yorodumi- EMDB-4747: CryoEM structure of calcium-bound human TMEM16K / Anoctamin 10 in... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-4747 | |||||||||
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Title | CryoEM structure of calcium-bound human TMEM16K / Anoctamin 10 in detergent (low Ca2+, closed form) | |||||||||
Map data | RELION3 postprocessed map sharpened with a bfactor of -176A**2 | |||||||||
Sample |
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Function / homology | Function and homology information intracellularly calcium-gated chloride channel activity / calcium-activated cation channel activity / chloride transport / chloride channel activity / monoatomic cation transport / chloride transmembrane transport / monoatomic ion transmembrane transport / Stimuli-sensing channels / Induction of Cell-Cell Fusion / intracellular membrane-bounded organelle ...intracellularly calcium-gated chloride channel activity / calcium-activated cation channel activity / chloride transport / chloride channel activity / monoatomic cation transport / chloride transmembrane transport / monoatomic ion transmembrane transport / Stimuli-sensing channels / Induction of Cell-Cell Fusion / intracellular membrane-bounded organelle / membrane / plasma membrane Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.2 Å | |||||||||
Authors | Pike ACW / Bushell SR / Shintre CA / Tessitore A / Chu A / Mukhopadhyay S / Shrestha L / Chalk R / Burgess-Brown NA / Love J ...Pike ACW / Bushell SR / Shintre CA / Tessitore A / Chu A / Mukhopadhyay S / Shrestha L / Chalk R / Burgess-Brown NA / Love J / Huiskonen JT / Edwards AM / Arrowsmith CH / Bountra C / Carpenter EP / Structural Genomics Consortium (SGC) | |||||||||
Funding support | United Kingdom, 2 items
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Citation | Journal: Nat Commun / Year: 2019 Title: The structural basis of lipid scrambling and inactivation in the endoplasmic reticulum scramblase TMEM16K. Authors: Simon R Bushell / Ashley C W Pike / Maria E Falzone / Nils J G Rorsman / Chau M Ta / Robin A Corey / Thomas D Newport / John C Christianson / Lara F Scofano / Chitra A Shintre / Annamaria ...Authors: Simon R Bushell / Ashley C W Pike / Maria E Falzone / Nils J G Rorsman / Chau M Ta / Robin A Corey / Thomas D Newport / John C Christianson / Lara F Scofano / Chitra A Shintre / Annamaria Tessitore / Amy Chu / Qinrui Wang / Leela Shrestha / Shubhashish M M Mukhopadhyay / James D Love / Nicola A Burgess-Brown / Rebecca Sitsapesan / Phillip J Stansfeld / Juha T Huiskonen / Paolo Tammaro / Alessio Accardi / Elisabeth P Carpenter / Abstract: Membranes in cells have defined distributions of lipids in each leaflet, controlled by lipid scramblases and flip/floppases. However, for some intracellular membranes such as the endoplasmic ...Membranes in cells have defined distributions of lipids in each leaflet, controlled by lipid scramblases and flip/floppases. However, for some intracellular membranes such as the endoplasmic reticulum (ER) the scramblases have not been identified. Members of the TMEM16 family have either lipid scramblase or chloride channel activity. Although TMEM16K is widely distributed and associated with the neurological disorder autosomal recessive spinocerebellar ataxia type 10 (SCAR10), its location in cells, function and structure are largely uncharacterised. Here we show that TMEM16K is an ER-resident lipid scramblase with a requirement for short chain lipids and calcium for robust activity. Crystal structures of TMEM16K show a scramblase fold, with an open lipid transporting groove. Additional cryo-EM structures reveal extensive conformational changes from the cytoplasmic to the ER side of the membrane, giving a state with a closed lipid permeation pathway. Molecular dynamics simulations showed that the open-groove conformation is necessary for scramblase activity. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_4747.map.gz | 41.6 MB | EMDB map data format | |
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Header (meta data) | emd-4747-v30.xml emd-4747.xml | 22.3 KB 22.3 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_4747_fsc.xml | 8.2 KB | Display | FSC data file |
Images | emd_4747.png | 178.8 KB | ||
Masks | emd_4747_msk_1.map | 45.2 MB | Mask map | |
Others | emd_4747_half_map_1.map.gz emd_4747_half_map_2.map.gz | 33.7 MB 33.7 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-4747 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-4747 | HTTPS FTP |
-Related structure data
Related structure data | 6r7yMC 4746C 4748C 5oc9C 6r65C 6r7xC 6r7zC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_4747.map.gz / Format: CCP4 / Size: 45.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | RELION3 postprocessed map sharpened with a bfactor of -176A**2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.822 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_4747_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: RELION3 halfmap1
File | emd_4747_half_map_1.map | ||||||||||||
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Annotation | RELION3 halfmap1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: RELION3 halfmap2
File | emd_4747_half_map_2.map | ||||||||||||
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Annotation | RELION3 halfmap2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Anoctamin 10
Entire | Name: Anoctamin 10Calcium-dependent chloride channel |
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Components |
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-Supramolecule #1: Anoctamin 10
Supramolecule | Name: Anoctamin 10 / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 153 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) / Recombinant plasmid: PFB-CT10HF-LIC |
-Macromolecule #1: Anoctamin-10
Macromolecule | Name: Anoctamin-10 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 77.276016 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MKVTLSALDT SESSFTPLVV IELAQDVKEE TKEWLKNRII AKKKDGGAQL LFRPLLNKYE QETLENQNLY LVGASKIRML LGAEAVGLV KECNDNTMRA FTYRTRQNFK GFDDNNDDFL TMAECQFIIK HELENLRAKD EKMIPGYPQA KLYPGKSLLR R LLTSGIVI ...String: MKVTLSALDT SESSFTPLVV IELAQDVKEE TKEWLKNRII AKKKDGGAQL LFRPLLNKYE QETLENQNLY LVGASKIRML LGAEAVGLV KECNDNTMRA FTYRTRQNFK GFDDNNDDFL TMAECQFIIK HELENLRAKD EKMIPGYPQA KLYPGKSLLR R LLTSGIVI QVFPLHDSEA LKKLEDTWYT RFALKYQPID SIRGYFGETI ALYFGFLEYF TFALIPMAVI GLPYYLFVWE DY DKYVIFA SFNLIWSTVI LELWKRGCAN MTYRWGTLLM KRKFEEPRPG FHGVLGINSI TGKEEPLYPS YKRQLRIYLV SLP FVCLCL YFSLYVMMIY FDMEVWALGL HENSGSEWTS VLLYVPSIIY AIVIEIMNRL YRYAAEFLTS WENHRLESAY QNHL ILKVL VFNFLNCFAS LFYIAFVLKD MKLLRQSLAT LLITSQILNQ IMESFLPYWL QRKHGVRVKR KVQALKADID ATLYE QVIL EKEMGTYLGT FDDYLELFLQ FGYVSLFSCV YPLAAAFAVL NNFTEVNSDA LKMCRVFKRP FSEPSANIGV WQLAFE TMS VISVVTNCAL IGMSPQVNAV FPESKADLIL IVVAVEHALL ALKFILAFAI PDKPRHIQMK LARLEFESLE ALKQQQM KL VTENLKEEPM ESGKEKATAE NLYFQ |
-Macromolecule #2: CALCIUM ION
Macromolecule | Name: CALCIUM ION / type: ligand / ID: 2 / Number of copies: 6 / Formula: CA |
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Molecular weight | Theoretical: 40.078 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 3 mg/mL | |||||||||||||||
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Buffer | pH: 7.5 Component:
Details: No additional calcium added. Background levels of calcium estimated to be ca. 500nM | |||||||||||||||
Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR | |||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV / Details: blot for 4.5sec prior to plunging. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.75 µm / Nominal defocus min: 1.0 µm |
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-20 / Number grids imaged: 1 / Number real images: 1975 / Average exposure time: 8.0 sec. / Average electron dose: 56.5 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
-Atomic model buiding 1
Details | Refined using phenix.real_space_refine with NCS constraints and model reference restraints used (PDB id: TBC) against RELION3 post-processed map. |
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Refinement | Space: REAL / Protocol: AB INITIO MODEL / Overall B value: 176 / Target criteria: Correlation coef |
Output model | PDB-6r7y: |