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Electron cryomicroscopy reveals different F1+F2 protein States in intact parainfluenza virions.

by single particle reconstruction, at 15 A resolution

Movie

Orientation:

#1: Surface view with section colored by density value, Surface level: -100, Image by UCSF CHIMERA

#2: Surface view colored by cylindrical radius, Surface level: -100, Image by UCSF CHIMERA

Entry
Summary
Database / IDEM DATA BANK (EMDB) / 1421
TitleElectron cryomicroscopy reveals different F1+F2 protein States in intact parainfluenza virions.
MapIn-situ 3D reconstruction of the ectodomain of the PIV5 F protein determined from cryo-negative stain electron micrographs
SampleF-Protein in intact Parainfluenzavirus 5
AuthorsLudwig K, Schade B, Boettcher C, Korte T
DateDeposition: 2007-09-07, Header release: 2007-09-11, Map release: 2008-04-07, Last update: 2012-10-24
EMDB SitesEMDB @PDBe (EU), EMDB @RCSB (USA)
Structure Visualization
MoviesMovie Page

#1: Surface view with section colored by density value, Surface level: -100, Image by UCSF CHIMERA

#2: Surface view colored by cylindrical radius, Surface level: -100, Image by UCSF CHIMERA

Supplemental images
Structure viewersYorodumi, Launch PeppeR (About PeppeR), Volume viewer (RCSB, PDBe)
Related Structure Data
Article
Citation - Primary
ArticleJ. Virol., Vol. 82, Issue 7, Page 3775-81, Year 2008
TitleElectron cryomicroscopy reveals different F1+F2 protein States in intact parainfluenza virions.
AuthorsKai Ludwig, Boris Schade, Christoph Böttcher, Thomas Korte, Nina Ohlwein, Bolormaa Baljinnyam, Michael Veit, Andreas Herrmann
Forschungszentrum für Elektronenmikroskopie, Freie Universität Berlin, Fabeckstr. 36a, D-14195 Berlin, Germany.
KeywordsAnimals, Cattle, Cell Fusion, Cell Line, Chick Embryo, Cryoelectron Microscopy, Image Processing, Computer-Assisted, Paramyxoviridae (ultrastructure), Protein Structure, Tertiary, Viral Proteins (ultrastructure), Virion (ultrastructure)
LinksDOI: 10.1128/JVI.02154-07, PubMed: 18216117, PMC: PMC2268498
Map
Fileemd_1421.map.gz ( map file in CCP4 format, 1301 KB )
Projections & SlicesSize of images:
AxesZ (Sec.)Y (Row.)X (Col.)
110 pix
1.56 A/pix
= 171.6 A
110 pix
1.56 A/pix
= 171.6 A
110 pix
1.56 A/pix
= 171.6 A

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Density
Contour Level:-95 (by emdb), -100 (movie #1):
Minimum - Maximum: -128 - 122
Average (Standard dev.): -124.687 (18.0757)
Data TypeEnvelope stored as signed bytes
Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions110110110
Origin-55-54-55
Limit545554
Spacing110110110
Unit CellA= B= C: 171.6 A
Alpha=beta=gamma: 90 degrees
Pixel SpacingX= Y= Z: 1.56 A
CCP4 map header info
modeenvelope stored as signed bytes (from -128 lowest to 127 highest)
A/pix X/Y/Z1.561.561.56
M x/y/z110110110
origin x/y/z0.0000.0000.000
length x/y/z171.600171.600171.600
alpha/beta/gamma90.00090.00090.000
start NX/NY/NZ-30-30-49
NX/NY/NZ6060100
MAP C/R/S123
start NC/NR/NS-54-55-55
NC/NR/NS110110110
start NC,NX/NR,NY/NS,NZ
NC,NX/NR,NY/NS,NZ
D min/max/mean-128.000122.000-124.687
Annotation DetailsIn-situ 3D reconstruction of the ectodomain of the PIV5 F protein determined from cryo-negative stain electron micrographs
Supplement
Images
Images
Sample
NameF-Protein in intact Parainfluenzavirus 5
Number of Components1
Oligomeric StateF-Protein Homotrimer
Theoretical Mass0.15MDa
Detailssample of fusion active virions (proven by fluorescence spectroscopy)of PIV5 (strain W3A)
Component #1: protein - F-Protein
Scientific nameF-Protein
Experimental Mass0.15 MDa
Oligomeric DetailsHomotrimer
Scientific Name of SpeciesParainfluenza virus 5
NCBI taxonomy11207
StrainW3A
Recombinant expressionNo
Natural SourceCell Location: viral membrane
Experiment
Sample Preparation
Staining30 seconds absorption 60 seconds staining (1% phospho-tungstic acid, pH 7.4) vitrified in liquid ethane
Specimen Support Details200 mesh carbon coated collodium-supported copper grids
Specimen Stateparticle
BufferDetails: PBS (150 mM NaCl, 5.8 mM NaH2PO4/Na2HPO4)
pH: 7.4
Vitrification
Cryogen NameETHANE
InstrumentHOMEMADE PLUNGER
MethodA small vial of ethane is placed inside a larger liquid nitrogen reservoir. The grid holding a few microliters of the sample is held in place at the bottom of a plunger by the means of fine tweezers. Once the ethane in the vial is completely frozen, it needs to be slightly melted. When the liquid ethane is ready, a piece of filter paper is then pressed against the sample to blot of excess buffer, sufficient to leave a thin layer on the grid. After a redetermined time, the filter paper is removed, and the plunger is allowed to drop into the liquid ethane. Once the grid enters the liquid ethane, the sample is rapidly frozen,and the grid is transferred under liquid nitrogen to a storage box immersed liquid nitrogen for later use in the microscope.
DetailsVitrification instrument: self-construction
Imaging
MicroscopeFEI TECNAI F20
Electron Gun
Electron SourceFIELD EMISSION GUN
Accelerating Voltage160 kV
Electron Dose12 e/A**2
Illumination ModeFLOOD BEAM
Lens
MagnificationNominal: 50000, Calibrated: 51064
Astigmatismobjective lens astigmatism was corrected at 100,000 times magnification
Nominal Cs2 mm
Imaging ModeBRIGHT FIELD
Defocus1501 nm - 2066 nm
Specimen Holder
HolderSide entry liquid nitrogen-cooled cryo specimen holder
ModelGATAN LIQUID NITROGEN
Temperature92 K
Camera
DetectorKODAK SO-163 FILM
Image Acquisition
#1
Number of Digital Images175
Sampling Size4
Quant Bit Number8
ScannerPRIMESCAN
Processing
Methodsingle particle reconstruction
3D reconstruction
AlgorithmCommon lines
SoftwareImagic
CTF CorrectionMSA-based
Resolution By Author15 A
Resolution MethodFSC 3 SIGMA
Single Particle
Number of Projections5700
Detailswell resolved spike-like proteins protruding from the viral membrane suitable for single particle analysis were interactively selected
Applied SymmetryC3 (3 fold cyclic)
Download
Data from EMDB
Header (meta data in XML format)emd-1421.xml (8.5 KB)
Map dataemd_1421.map.gz (75.5 KB)
Images1421.gif (43.5 KB)
emd_1421.tif (258.8 KB)
FTP directoryftp://ftp.pdbj.org/pub/emdb/structures/EMD-1421
Movie files
movie #1
.mp4 (H.264/MPEG-4 AVC format), 3.5 MB
.webm (WebM/VP8 format), 4.5 MB
Session file for UCSF-Chimera, 26.3 KB
movie #2
.mp4 (H.264/MPEG-4 AVC format), 3.2 MB
.webm (WebM/VP8 format), 4.1 MB
Session file for UCSF-Chimera, 26.3 KB