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- EMDB-2531: Tomographic subvolume average of EFF-1 fusogen on extracellular v... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-2531 | |||||||||
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Title | Tomographic subvolume average of EFF-1 fusogen on extracellular vesicles | |||||||||
![]() | Segmented membrane structure from EFF-1 coated vesicles | |||||||||
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![]() | cell-cell fusion / extracellular fusion / membrane fusion / fusogen / pre-fusion state | |||||||||
Biological species | ![]() | |||||||||
Method | subtomogram averaging / cryo EM | |||||||||
![]() | Zeev-Ben-Mordehai T / Vasishtan D / Siebert CA / Grunewald K | |||||||||
![]() | ![]() Title: The full-length cell-cell fusogen EFF-1 is monomeric and upright on the membrane. Authors: Tzviya Zeev-Ben-Mordehai / Daven Vasishtan / C Alistair Siebert / Kay Grünewald / ![]() Abstract: Fusogens are membrane proteins that remodel lipid bilayers to facilitate membrane merging. Although several fusogen ectodomain structures have been solved, structural information on full-length, ...Fusogens are membrane proteins that remodel lipid bilayers to facilitate membrane merging. Although several fusogen ectodomain structures have been solved, structural information on full-length, natively membrane-anchored fusogens is scarce. Here we present the electron cryo microscopy three-dimensional reconstruction of the Caenorhabditis elegans epithelial fusion failure 1 (EFF-1) protein natively anchored in cell-derived membrane vesicles. This reveals a membrane protruding, asymmetric, elongated monomer. Flexible fitting of a protomer of the EFF-1 crystal structure, which is homologous to viral class-II fusion proteins, shows that EFF-1 has a hairpin monomeric conformation before fusion. These structural insights, when combined with our observations of membrane-merging intermediates between vesicles, enable us to propose a model for EFF-1 mediated fusion. This process, involving identical proteins on both membranes to be fused, follows a mechanism that shares features of SNARE-mediated fusion while using the structural building blocks of the unilaterally acting class-II viral fusion proteins. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 224 KB | ![]() | |
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Header (meta data) | ![]() ![]() | 9.5 KB 9.5 KB | Display Display | ![]() |
Images | ![]() | 70.6 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 195.2 KB | Display | ![]() |
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Full document | ![]() | 194.3 KB | Display | |
Data in XML | ![]() | 4.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
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Links
EMDB pages | ![]() ![]() |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Segmented membrane structure from EFF-1 coated vesicles | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 3.8 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : Segmented membrane from epithelial fusion failure 1 (EFF-1) Isofo...
Entire | Name: Segmented membrane from epithelial fusion failure 1 (EFF-1) Isoform A on extracellular vesicles |
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Components |
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-Supramolecule #1000: Segmented membrane from epithelial fusion failure 1 (EFF-1) Isofo...
Supramolecule | Name: Segmented membrane from epithelial fusion failure 1 (EFF-1) Isoform A on extracellular vesicles type: sample / ID: 1000 / Number unique components: 1 |
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-Supramolecule #1: Cell-derived vesicle membrane
Supramolecule | Name: Cell-derived vesicle membrane / type: organelle_or_cellular_component / ID: 1 Details: Membrane structure of EFF-1 coated extracellular vesicles Recombinant expression: No |
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Source (natural) | Organism: ![]() |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | subtomogram averaging |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.4 / Details: 25mM HEPES, 130mM NaCl |
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Grid | Details: Holey carbon on top of 200 mesh gold grid. |
Vitrification | Cryogen name: ETHANE-PROPANE MIXTURE / Chamber temperature: 77 K / Instrument: HOMEMADE PLUNGER / Method: Blot for 3 seconds before plunging |
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Electron microscopy
Microscope | FEI POLARA 300 |
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Temperature | Min: 80 K / Max: 100 K / Average: 85 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 115,000 times magnification Legacy - Electron beam tilt params: +1 to -1 mradians |
Specialist optics | Energy filter - Name: Gatan Quantum 964 / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV |
Date | Mar 3, 2013 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Digitization - Sampling interval: 30 µm / Number real images: 78 / Average electron dose: 60 e/Å2 / Details: 2 tomograms each created from 39 projection images / Bits/pixel: 16 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 78950 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 95000 |
Sample stage | Specimen holder: Liquid nitrogen cooled / Specimen holder model: GATAN HELIUM / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 ° |
Experimental equipment | ![]() Model: Tecnai Polara / Image courtesy: FEI Company |
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Image processing
Details | Subtomograms were selected using a semi-automated picking algorithm |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Software - Name: PEET / Number subtomograms used: 1973 |
Final 3D classification | Number classes: 1 |