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Cryo-electron tomography reveals novel interactions and doublet-specific structures in the I1 dynein

by subtomogram averaging, at 39 A resolution

Movie

Orientation:

#1: Surface view with section colored by density value, Surface level: 127.5, Image by UCSF CHIMERA

#2: Surface view colored by cylindrical radius, Surface level: 127.5, Image by UCSF CHIMERA

Entry
Summary
Database / IDEM DATA BANK (EMDB) / 5330
TitleCryo-electron tomography reveals novel interactions and doublet-specific structures in the I1 dynein
MapThis is a subtomogram average of the I1 inner dynein complex in wild type Chlamydomonas flagella
SampleCryo-electron tomography and subtomographic average (750 axonemal repeats) of isolated axonemes of wild type Chlamydomonas (CC 125, 137c), I1 dynein complex (dynein f) is bound to the doublet microtubule and is connected to neighboring structures.
Keywordsaxoneme, molecular motor, motility regulation, flagella
AuthorsHeuser T, Barber CF, Lin J, Krell J, Rebesco M, Porter ME, Nicastro D
DateDeposition: 2011-08-10, Header release: 2011-08-11, Map release: 2012-06-21, Last update: 2014-04-02
EMDB SitesEMDB @PDBe (EU), EMDB @RCSB (USA)
Structure Visualization
MoviesMovie Page

#1: Surface view with section colored by density value, Surface level: 127.5, Image by UCSF CHIMERA

#2: Surface view colored by cylindrical radius, Surface level: 127.5, Image by UCSF CHIMERA

Supplemental images
Structure viewersYorodumi, Launch PeppeR (About PeppeR), Volume viewer (RCSB, PDBe)
Related Structure Data
Similar strucutres (beta)
List of similar structure data about Omokage system
Article
Citation - Primary
ArticleProc. Natl. Acad. Sci. U.S.A., Vol. 109, Issue 30, Page E2067-76, Year 2012
TitleCryoelectron tomography reveals doublet-specific structures and unique interactions in the I1 dynein.
AuthorsThomas Heuser, Cynthia F Barber, Jianfeng Lin, Jeremy Krell, Matthew Rebesco, Mary E Porter, Daniela Nicastro
Biology Department, Rosenstiel Center, Brandeis University, Waltham, MA 02454, USA.
KeywordsAxoneme (chemistry), Chlamydomonas (chemistry), Cryoelectron Microscopy (methods), Dyneins (chemistry, 3.6.4.2), Image Processing, Computer-Assisted, Immunoblotting, Microtubules (chemistry), Mutation (genetics), Proteomics, Signal Transduction (genetics)
LinksDOI: 10.1073/pnas.1120690109, PubMed: 22733763, PMC: PMC3409752
Map
Fileemd_5330.map.gz ( map file in CCP4 format, 345 KB )
Projections & SlicesSize of images:
AxesZ (Sec.)Y (Row.)X (Col.)
40 pix
9.5 A/pix
= 380. A
40 pix
9.5 A/pix
= 522.5 A
55 pix
9.5 A/pix
= 380. A

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Density
Contour Level:127.5 (by author), 127.5 (movie #1):
Minimum - Maximum: 109.59519958 - 148.88063049
Average (Standard dev.): 124.20162964 (5.68234873)
Data TypeImage stored as Reals
Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions405540
Origin-53-21-64
Limit-1433-25
Spacing554040
Unit CellA: 380 A, B: 522.5 A, C: 380 A
Alpha=beta=gamma: 90 degrees
Pixel SpacingX= Y= Z: 9.5 A
CCP4 map header info
modeImage stored as Reals
A/pix X/Y/Z9.59.59.5
M x/y/z405540
origin x/y/z0.0000.0000.000
length x/y/z380.000522.500380.000
alpha/beta/gamma90.00090.00090.000
start NX/NY/NZ-207-207-206
NX/NY/NZ414414414
MAP C/R/S123
start NC/NR/NS-21-53-64
NC/NR/NS554040
start NC,NX/NR,NY/NS,NZ
NC,NX/NR,NY/NS,NZ
D min/max/mean109.595148.881124.202
Annotation DetailsThis is a subtomogram average of the I1 inner dynein complex in wild type Chlamydomonas flagella
Supplement
Images
Images
Sample
NameCryo-electron tomography and subtomographic average (750 axonemal repeats) of isolated axonemes of wild type Chlamydomonas (CC 125, 137c), I1 dynein complex (dynein f) is bound to the doublet microtubule and is connected to neighboring structures.
Number of Components1
Component #1: cellular-component - dynein f
Scientific nameI1 dynein complex
Common Namedynein f
Number of Copies2
Scientific Name of SpeciesChlamydomonas reinhardtii
Common Name of Speciesunicellular green algae
NCBI taxonomy3055
StrainCC-125, 137c
Recombinant expressionNo
Natural SourceOrganelle: eukaryotic flagella
Cell: Chlamydomonas reinhardtii
Experiment
Sample Preparation
Specimen Conc1 mg/ml
Specimen Support DetailsQuantifoil holey carbon grids Cu 200 mesh R2/2
BufferDetails: 10 mM HEPES, pH 7.4, 25 mM NaCl, 4 mM MgSO4, 1 mM EGTA, 0.1 mM EDTA
pH: 7.4
Vitrification
Methodfront-side blotting for 2-3 seconds
Cryogen NameETHANE
InstrumentHOMEMADE PLUNGER
Temperature100 Kelvin
Imaging
MicroscopeFEI TECNAI F30
Date20-FEB-2004
Electron Gun
Electron SourceFIELD EMISSION GUN
Accelerating Voltage300 kV
Electron Dose100 e/A**2
Illumination ModeOTHER
Lens
MagnificationNominal: 13500
Imaging ModeBRIGHT FIELD
Defocus6000 nm - 8000 nm
Energy FilterEnergy Filter: GATAN postcolumn filter GIF , Energy Window: 0-20 eV
Specimen Holder
HolderEucentric
ModelGATAN LIQUID NITROGEN
Tilt Angle-60 degrees - 60 degrees
Temperature80 K
Camera
DetectorGENERIC GATAN (2k x 2k)
Processing
Methodsubtomogram averaging
3D reconstruction
Algorithmfiducial alignment and weighted backprojection
SoftwareIMOD
DetailsFinal maps were calculated by averaging 750 particles from 5 tomograms
Resolution By Author39 A
Resolution MethodFSC 0.5
Subtomogram Averaging
Details750 axonemal repeats (96 nm long) from 5 tomograms (reconstructed using fiducial alignment and weighted backprojection, IMOD software, Kremer et al. 1996) were aligned and averaged using the PEET software (bio3d.colorado.edu, Nicastro et al. 2006). Average number of tilts used in the 3D reconstructions: 80. Average tomographic tilt angle increment: 1.5.
Download
Data from EMDB
Header (meta data in XML format)emd-5330.xml (7.6 KB)
Map dataemd_5330.map.gz (284.5 KB)
Imagesemd_5330_1.tif (184.9 KB)
FTP directoryftp://ftp.pdbj.org/pub/emdb/structures/EMD-5330
Movie files
movie #1
.mp4 (H.264/MPEG-4 AVC format), 3.3 MB
.webm (WebM/VP8 format), 5.4 MB
Session file for UCSF-Chimera, 26.6 KB
movie #2
.mp4 (H.264/MPEG-4 AVC format), 3 MB
.webm (WebM/VP8 format), 4.4 MB
Session file for UCSF-Chimera, 26.6 KB