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- PDB-6qm6: Cryo-EM structure of calcium-free nhTMEM16 lipid scramblase in DDM -
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Open data
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Basic information
Entry | Database: PDB / ID: 6qm6 | |||||||||
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Title | Cryo-EM structure of calcium-free nhTMEM16 lipid scramblase in DDM | |||||||||
![]() | Predicted protein | |||||||||
![]() | MEMBRANE PROTEIN / lipid scrambles / TMEM16 | |||||||||
Function / homology | ![]() cortical endoplasmic reticulum / chloride channel activity / identical protein binding / metal ion binding / plasma membrane Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å | |||||||||
![]() | Kalienkova, V. / Clerico Mosina, V. / Bryner, L. / Oostergetel, G.T. / Dutzler, R. / Paulino, C. | |||||||||
Funding support | ![]() ![]()
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![]() | ![]() Title: Stepwise activation mechanism of the scramblase nhTMEM16 revealed by cryo-EM. Authors: Valeria Kalienkova / Vanessa Clerico Mosina / Laura Bryner / Gert T Oostergetel / Raimund Dutzler / Cristina Paulino / ![]() ![]() Abstract: Scramblases catalyze the movement of lipids between both leaflets of a bilayer. Whereas the X-ray structure of the protein nhTMEM16 has previously revealed the architecture of a Ca-dependent lipid ...Scramblases catalyze the movement of lipids between both leaflets of a bilayer. Whereas the X-ray structure of the protein nhTMEM16 has previously revealed the architecture of a Ca-dependent lipid scramblase, its regulation mechanism has remained elusive. Here, we have used cryo-electron microscopy and functional assays to address this question. Ca-bound and Ca-free conformations of nhTMEM16 in detergent and lipid nanodiscs illustrate the interactions with its environment and they reveal the conformational changes underlying its activation. In this process, Ca binding induces a stepwise transition of the catalytic subunit cavity, converting a closed cavity that is shielded from the membrane in the absence of ligand, into a polar furrow that becomes accessible to lipid headgroups in the Ca-bound state. Additionally, our structures demonstrate how nhTMEM16 distorts the membrane at both entrances of the subunit cavity, thereby decreasing the energy barrier for lipid movement. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 241.7 KB | Display | ![]() |
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PDB format | ![]() | 195.9 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 4589MC ![]() 4587C ![]() 4588C ![]() 4592C ![]() 4593C ![]() 4594C ![]() 6qm4C ![]() 6qm5C ![]() 6qm9C ![]() 6qmaC ![]() 6qmbC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Components
#1: Protein | Mass: 83200.008 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: nhTMEM16 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Value: 0.166 MDa / Experimental value: NO |
Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 7.6 / Details: 5 mM Hepes 7.6, 150 mM NaCl, 0.03% DDM, 2 mM EGTA |
Specimen | Conc.: 3.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: at 5mA / Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 288 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 49407 X / Calibrated magnification: 49407 X / Nominal defocus max: 3000 nm / Nominal defocus min: 300 nm / Calibrated defocus min: 300 nm / Calibrated defocus max: 3000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 105 K / Temperature (min): 90 K |
Image recording | Average exposure time: 9 sec. / Electron dose: 52 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 9 / Num. of real images: 3351 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Width: 3838 / Height: 3710 / Movie frames/image: 60 / Used frames/image: 1-60 |
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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: 570203 | ||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 238070 / Algorithm: BACK PROJECTION / Num. of class averages: 9 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | Space: REAL |