+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-8344 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Influenza virus membrane fusion with liposomes | |||||||||
Map data | Influenza virus membrane fusion with liposomes | |||||||||
Sample |
| |||||||||
Biological species | Influenza A virus | |||||||||
Method | electron tomography / cryo EM | |||||||||
Authors | Gui L / Lee K | |||||||||
Funding support | United States, 1 items
| |||||||||
Citation | Journal: J Virol / Year: 2016 Title: Visualization and Sequencing of Membrane Remodeling Leading to Influenza Virus Fusion. Authors: Long Gui / Jamie L Ebner / Alexander Mileant / James A Williams / Kelly K Lee / Abstract: Protein-mediated membrane fusion is an essential step in many fundamental biological events, including enveloped virus infection. The nature of protein and membrane intermediates and the sequence of ...Protein-mediated membrane fusion is an essential step in many fundamental biological events, including enveloped virus infection. The nature of protein and membrane intermediates and the sequence of membrane remodeling during these essential processes remain poorly understood. Here we used cryo-electron tomography (cryo-ET) to image the interplay between influenza virus and vesicles with a range of lipid compositions. By following the population kinetics of membrane fusion intermediates imaged by cryo-ET, we found that membrane remodeling commenced with the hemagglutinin fusion protein spikes grappling onto the target membrane, followed by localized target membrane dimpling as local clusters of hemagglutinin started to undergo conformational refolding. The local dimples then transitioned to extended, tightly apposed contact zones where the two proximal membrane leaflets were in most cases indistinguishable from each other, suggesting significant dehydration and possible intermingling of the lipid head groups. Increasing the content of fusion-enhancing cholesterol or bis-monoacylglycerophosphate in the target membrane led to an increase in extended contact zone formation. Interestingly, hemifused intermediates were found to be extremely rare in the influenza virus fusion system studied here, most likely reflecting the instability of this state and its rapid conversion to postfusion complexes, which increased in population over time. By tracking the populations of fusion complexes over time, the architecture and sequence of membrane reorganization leading to efficient enveloped virus fusion were thus resolved. IMPORTANCE: Enveloped viruses employ specialized surface proteins to mediate fusion of cellular and viral membranes that results in the formation of pores through which the viral genetic material is ...IMPORTANCE: Enveloped viruses employ specialized surface proteins to mediate fusion of cellular and viral membranes that results in the formation of pores through which the viral genetic material is delivered to the cell. For influenza virus, the trimeric hemagglutinin (HA) glycoprotein spike mediates host cell attachment and membrane fusion. While structures of a subset of conformations and parts of the fusion machinery have been characterized, the nature and sequence of membrane deformations during fusion have largely eluded characterization. Building upon studies that focused on early stages of HA-mediated membrane remodeling, here cryo-electron tomography (cryo-ET) was used to image the three-dimensional organization of intact influenza virions at different stages of fusion with liposomes, leading all the way to completion of the fusion reaction. By monitoring the evolution of fusion intermediate populations over the course of acid-induced fusion, we identified the progression of membrane reorganization that leads to efficient fusion by an enveloped virus. | |||||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_8344.map.gz | 596.9 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-8344-v30.xml emd-8344.xml | 10.3 KB 10.3 KB | Display Display | EMDB header |
Images | emd_8344.png | 155.1 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-8344 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8344 | HTTPS FTP |
-Validation report
Summary document | emd_8344_validation.pdf.gz | 78.2 KB | Display | EMDB validaton report |
---|---|---|---|---|
Full document | emd_8344_full_validation.pdf.gz | 77.3 KB | Display | |
Data in XML | emd_8344_validation.xml.gz | 499 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8344 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8344 | HTTPS FTP |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|
-Map
File | Download / File: emd_8344.map.gz / Format: CCP4 / Size: 905.5 MB / Type: IMAGE STORED AS SIGNED BYTE | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Influenza virus membrane fusion with liposomes | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 3.19 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
|
-Supplemental data
-Sample components
-Entire : Influenza A virus
Entire | Name: Influenza A virus |
---|---|
Components |
|
-Supramolecule #1: Influenza A virus
Supramolecule | Name: Influenza A virus / type: virus / ID: 1 / Parent: 0 Details: X31 (H3N2) influenza A virus grown in embryonated chicken eggs was purchased from Charles River Lab. NCBI-ID: 11320 / Sci species name: Influenza A virus / Sci species strain: X31 (H3N2) / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: No |
---|
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | electron tomography |
Aggregation state | particle |
-Sample preparation
Concentration | 1.0 mg/mL | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Buffer | pH: 5.5 Component:
| ||||||||||||
Grid | Model: C-flat / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 25 K / Instrument: FEI VITROBOT MARK IV Details: 3 microliters of solution was added to glow-discharged holey carbon-coated grids (C-flat, 200 mesh, Electron Microscopy Sciences) and plunge-frozen in liquid ethane using an FEI Vitrobot Mark IV.. | ||||||||||||
Sectioning | Other: NO SECTIONING | ||||||||||||
Fiducial marker | Manufacturer: Electron Microscopy Sciences / Diameter: 10 nm |
-Electron microscopy
Microscope | FEI TECNAI F20 |
---|---|
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-11 / Average exposure time: 1.7 sec. / Average electron dose: 1.5 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 11500 |
Sample stage | Specimen holder model: GATAN 626 SINGLE TILT LIQUID NITROGEN CRYO TRANSFER HOLDER Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Tecnai F20 / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Algorithm: BACK PROJECTION / Software - Name: IMOD (ver. 4.7) / Number images used: 60 |
---|