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- EMDB-1561: Electron tomography of isometrically contracting insect flight muscle -

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Database: EMDB / ID: 1561
TitleElectron tomography of isometrically contracting insect flight muscle
Keywordsinsect / muscle / myosin / troponin / tropomyosin / actin / light chains / thin filament / thick filament / electron microscopy / image processing / isometric contraction / freezing / freeze substitution / microtomy / multivariate data analysis
SampleIsometrically contracting asynchronous insect flight muscle
SourceLethocerus indicus / arthropod / Water bug
Map dataThis is a dual axis tomogram of quick frozen, freeze This is a dual axis tomogram of quick frozen, freeze substituted, plastic embedded, thin sectioned insect flight muscle from Lethocerus sp.
Methodelectron tomography
AuthorsWu S / Liu J / Reedy MC / Tregear RT / Winkler H / Franzini-Armstrong C / Sasaki H / Lucaveche C / Goldman YE / Reedy MK / Taylor KA
CitationPLoS ONE, 2010, 5

primary. PLoS ONE, 2010, 5 StrPapers
Electron tomography of cryofixed, isometrically contracting insect flight muscle reveals novel actin-myosin interactions.
Shenping Wu / Jun Liu / Mary C Reedy / Richard T Tregear / Hanspeter Winkler / Clara Franzini-Armstrong / Hiroyuki Sasaki / Carmen Lucaveche / Yale E Goldman / Michael K Reedy / Kenneth A Taylor

1. J. Struct. Biol., 2009, 168, 485-502 StrPapers
Methods for identifying and averaging variable molecular conformations in tomograms of actively contracting insect flight muscle.
Shenping Wu / Jun Liu / Mary C Reedy / Hanspeter Winkler / Michael K Reedy / Kenneth A Taylor

Validation ReportPDB-ID: 2w49

SummaryFull report
PDB-ID: 2w4a

SummaryFull report
PDB-ID: 2w4t

SummaryFull report
About validation report
DateDeposition: Sep 26, 2008 / Header (metadata) release: Sep 29, 2008 / Map release: Apr 12, 2010 / Last update: Sep 26, 2008

Structure visualization

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Supplemental images

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Fileemd_1561.map.gz (map file in CCP4 format, 588001 KB)
Projections & slices

Image control

AxesZ (Sec.)Y (Row.)X (Col.)
70 pix
6.9 Å/pix.
= 483. Å
1344 pix
6.9 Å/pix.
= 11040. Å
1600 pix
6.9 Å/pix.
= 9273.601 Å


Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 6.9 Å
Minimum - Maximum-3296.96 - 4848.86
Average (Standard dev.)5.83993 (532.04)


Space Group Number1
Map Geometry
Axis orderXYZ
CellA: 9273.6 Å / B: 11040 Å / C: 483 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z6.90000074404766.96.9
M x/y/z1344160070
origin x/y/z0.0000.0000.000
length x/y/z9273.60111040.000483.000
start NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
D min/max/mean-3296.9594848.8645.840

Supplemental data

Sample components

Entire Isometrically contracting asynchronous insect flight muscle

EntireName: Isometrically contracting asynchronous insect flight muscle
Details: This specimen is obtained from a quick frozen, isometrically contracting asynchronous insect flight muscle that has been freeze substituted, plastic embedded, and thin sectioned
Number of components: 8 / Oligomeric State: tissue

Component #1: cellular-component, myofibril

Cellular-componentName: myofibril / a.k.a: muscle / Oligomeric Details: muscle fibril
Details: The sample was enblock stained using uranyl acetate and tannic acid, and post stained with lead citrate and potassium permanganate
Recombinant expression: No
SourceSpecies: Lethocerus indicus / arthropod / Water bug
Source (natural)Organelle: myofibril / Location in cell: myooplasm
Organ or tissue: asynchronous dorsal longitudinal flight muscle

Experimental details

Sample preparation

Sample solutionBuffer solution: 20 mM MOPS buffer, 5 mM NaN3, and MgCl2, ATP, CaCl2, and EGTA in varying millimolar concentrations
StainingFreeze slammed fibers were freeze-substituted in acetone using a tannic aciduranyl acetate sequence, and ultimately embedded in Araldite-506 for thin-section electron microscopy. Ultrathin (25-30 nm) longitudinal sections were stained by permanganate-lead.
VitrificationInstrument: HOMEMADE PLUNGER / Cryogen name: HELIUM / Temperature: 4.5 K
Method: smash against a liquid helium cooled Au-coated Cu mirror
Details: Vitrification instrument: modified Heuser Cryopress freezing head

Electron microscopy imaging

ImagingMicroscope: FEI/PHILIPS CM300FEG/T
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
LensCs: 2 mm / Imaging mode: BRIGHT FIELD
Specimen HolderHolder: Gatan model 670 Ultrahigh tilt analytical holder / Model: OTHER / Tilt Angle: -72 - 72 deg.
CameraDetector: TVIPS TEMCAM-F224 (2k x 2k)

Image acquisition

Image acquisitionBit depth: 16
Details: Images recorded on a TVIPS Tem-Cam F224 2k x 2k CCD camera

Image processing

ProcessingMethod: electron tomography / Number of sections: 70
3D reconstructionAlgorithm: marker free alignment / Software: PROTOMO, IMOD
Details: Two tilt series at 90 degrees to one another were first independently aligned using marker-free alignment and area matching. The two resulting tomograms were then merged by patch correlation and volume warp using IMOD. Tomogram computed using weighted back projection.
Resolution method: FSC 0.5

Atomic model buiding

Output model

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