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Yorodumi- EMDB-1966: CFTR map generated from 2D crystals grown using the epitaxial method. -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-1966 | |||||||||
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Title | CFTR map generated from 2D crystals grown using the epitaxial method. | |||||||||
Map data | EM map generated from 2D crystals of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR or ABCC7). | |||||||||
Sample |
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Keywords | Cystic fibrosis / CFTR / ion channel / ATP binding cassette protein | |||||||||
Function / homology | Function and homology information ATPase-coupled lipid transmembrane transporter activity / Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate / ABC-type transporter activity / membrane => GO:0016020 / ATP hydrolysis activity / ATP binding / plasma membrane Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | electron crystallography / cryo EM / negative staining / Resolution: 9.0 Å | |||||||||
Authors | Rosenberg MF / ORyan LP / Hughes G / Zhao Z / Aleksandrov LA / Riordan JR / Ford RC | |||||||||
Citation | Journal: J Biol Chem / Year: 2011 Title: The cystic fibrosis transmembrane conductance regulator (CFTR): three-dimensional structure and localization of a channel gate. Authors: Mark F Rosenberg / Liam P O'Ryan / Guy Hughes / Zhefeng Zhao / Luba A Aleksandrov / John R Riordan / Robert C Ford / Abstract: Cystic fibrosis affects about 1 in 2500 live births and involves loss of transmembrane chloride flux due to a lack of a membrane protein channel termed the cystic fibrosis transmembrane conductance ...Cystic fibrosis affects about 1 in 2500 live births and involves loss of transmembrane chloride flux due to a lack of a membrane protein channel termed the cystic fibrosis transmembrane conductance regulator (CFTR). We have studied CFTR structure by electron crystallography. The data were compared with existing structures of other ATP-binding cassette transporters. The protein was crystallized in the outward facing state and resembled the well characterized Sav1866 transporter. We identified regions in the CFTR map, not accounted for by Sav1866, which were potential locations for the regulatory region as well as the channel gate. In this analysis, we were aided by the fact that the unit cell was composed of two molecules not related by crystallographic symmetry. We also identified regions in the fitted Sav1866 model that were missing from the map, hence regions that were either disordered in CFTR or differently organized compared with Sav1866. Apart from the N and C termini, this indicated that in CFTR, the cytoplasmic end of transmembrane helix 5/11 and its associated loop could be partly disordered (or alternatively located). | |||||||||
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_1966.map.gz | 7.4 MB | EMDB map data format | |
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Header (meta data) | emd-1966-v30.xml emd-1966.xml | 12.6 KB 12.6 KB | Display Display | EMDB header |
Images | 1966.png | 200.1 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-1966 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-1966 | HTTPS FTP |
-Validation report
Summary document | emd_1966_validation.pdf.gz | 222.4 KB | Display | EMDB validaton report |
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Full document | emd_1966_full_validation.pdf.gz | 221.5 KB | Display | |
Data in XML | emd_1966_validation.xml.gz | 4.9 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1966 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1966 | HTTPS FTP |
-Related structure data
Related structure data | 4a82MC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_1966.map.gz / Format: CCP4 / Size: 15.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | EM map generated from 2D crystals of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR or ABCC7). | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X: 0.7 Å / Y: 0.76 Å / Z: 2.5 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Cystic fibrosis transmembrane conductance regulator
Entire | Name: Cystic fibrosis transmembrane conductance regulator |
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Components |
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-Supramolecule #1000: Cystic fibrosis transmembrane conductance regulator
Supramolecule | Name: Cystic fibrosis transmembrane conductance regulator / type: sample / ID: 1000 Details: Human ortholog. Sample was glycosylated and mostly dephosphorylated. No nucleotide was added. Oligomeric state: 2 Monomers / Number unique components: 1 |
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Molecular weight | Experimental: 200 KDa / Theoretical: 168.142 KDa / Method: SDS-PAGE |
-Macromolecule #1: Cystic fibrosis transmembrane conductance regulator
Macromolecule | Name: Cystic fibrosis transmembrane conductance regulator / type: protein_or_peptide / ID: 1 / Name.synonym: CFTR Details: Human protein is glycosylated but non-phosphorylated, as expressed and purified from BHK cells. No nucleotide is present during purification and crystallisation. Number of copies: 2 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human / Cell: BHK |
Molecular weight | Experimental: 200 KDa / Theoretical: 168 KDa |
Recombinant expression | Organism: Chinese Hamster Kidney cells |
Sequence | GO: membrane => GO:0016020 InterPro: Cystic fibrosis transmembrane conductance regulator |
-Experimental details
-Structure determination
Method | negative staining, cryo EM |
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Processing | electron crystallography |
Aggregation state | 2D array |
-Sample preparation
Concentration | 0.05 mg/mL |
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Buffer | pH: 8 Details: 0.05% dodecyl maltoside, polyethylene glycol 4000 (10% w/v), 100mM ammonium sulphate, 50 mM Tris-HCl. |
Staining | Type: NEGATIVE / Details: Unstained |
Grid | Details: 400 mesh carbon, gold |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 80 K / Instrument: OTHER / Details: Vitrification instrument: Vitrobot / Method: 10 seconds |
Details | on carbon (epitaxial) |
Crystal formation | Details: on carbon (epitaxial) |
-Electron microscopy
Microscope | FEI POLARA 300 |
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Temperature | Min: 97 K / Max: 97 K / Average: 97 K |
Image recording | Category: CCD / Film or detector model: GENERIC GATAN (4k x 4k) / Digitization - Sampling interval: 1.9 µm / Number real images: 121 / Bits/pixel: 16 |
Tilt angle min | 0 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.1 µm |
Sample stage | Specimen holder: Eucentric / Specimen holder model: GATAN HELIUM / Tilt angle max: 70 / Tilt series - Axis1 - Min angle: 0 ° / Tilt series - Axis1 - Max angle: 70 ° |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 9.0 Å / Resolution method: OTHER / Software - Name: MRC Details: Lattice unbending and origin refinement were carried out with 2dx and 2dxmerge software. |
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Crystal parameters | Unit cell - A: 72.300 Å / Unit cell - B: 75.800 Å / Unit cell - C: 300.000 Å / Unit cell - γ: 125.00 ° / Unit cell - α: 90.00 ° / Unit cell - β: 90.00 ° / Plane group: P 1 |
CTF correction | Details: TTREFINE |
-Atomic model buiding 1
Initial model | PDB ID: |
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Software | Name: Chimera |
Details | Protocol: Rigid Body. Initial manual docking then refinement using Chimera fir-in-map routine. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT Target criteria: Cross-correlation between calculated map vs experimental map |
Output model | PDB-4a82: |