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- EMDB-1561: Electron tomography of isometrically contracting insect flight muscle -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-1561 | |||||||||
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Title | Electron tomography of isometrically contracting insect flight muscle | |||||||||
![]() | This 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. | |||||||||
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![]() | insect / muscle / myosin / troponin / tropomyosin / actin / light chains / thin filament / thick filament / electron microscopy / image processing / isometric contraction / freezing / freeze substitution / microtomy / multivariate data analysis | |||||||||
Function / homology | ![]() cytoskeletal motor regulator activity / troponin C binding / contractile muscle fiber / mitotic actomyosin contractile ring / mitotic actomyosin contractile ring contraction / troponin complex / Striated Muscle Contraction / regulation of muscle contraction / muscle myosin complex / myosin filament ...cytoskeletal motor regulator activity / troponin C binding / contractile muscle fiber / mitotic actomyosin contractile ring / mitotic actomyosin contractile ring contraction / troponin complex / Striated Muscle Contraction / regulation of muscle contraction / muscle myosin complex / myosin filament / actomyosin structure organization / locomotion / myosin complex / cytoskeletal motor activator activity / myosin II complex / microfilament motor activity / myofibril / sarcomere organization / tropomyosin binding / mesenchyme migration / troponin I binding / myosin heavy chain binding / filamentous actin / actin filament bundle / skeletal muscle thin filament assembly / striated muscle thin filament / actin filament bundle assembly / skeletal muscle contraction / skeletal muscle myofibril / mitotic cytokinesis / actin monomer binding / skeletal muscle fiber development / cardiac muscle contraction / stress fiber / skeletal muscle tissue development / titin binding / actin filament polymerization / post-embryonic development / filopodium / muscle contraction / actin filament / actin filament organization / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / calcium-dependent protein binding / actin filament binding / actin cytoskeleton / lamellipodium / actin binding / cell body / hydrolase activity / calmodulin binding / protein heterodimerization activity / protein domain specific binding / calcium ion binding / positive regulation of gene expression / magnesium ion binding / protein homodimerization activity / ATP binding / identical protein binding / cytoplasm Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | electron tomography / cryo EM / negative staining | |||||||||
![]() | Wu S / Liu J / Reedy MC / Tregear RT / Winkler H / Franzini-Armstrong C / Sasaki H / Lucaveche C / Goldman YE / Reedy MK / Taylor KA | |||||||||
![]() | Journal: J Struct Biol / Year: 2009 Title: Methods for identifying and averaging variable molecular conformations in tomograms of actively contracting insect flight muscle. Authors: Shenping Wu / Jun Liu / Mary C Reedy / Hanspeter Winkler / Michael K Reedy / Kenneth A Taylor / ![]() Abstract: During active muscle contraction, tension is generated through many simultaneous, independent interactions between the molecular motor myosin and the actin filaments. The ensemble of myosin motors ...During active muscle contraction, tension is generated through many simultaneous, independent interactions between the molecular motor myosin and the actin filaments. The ensemble of myosin motors displays heterogeneous conformations reflecting different mechanochemical steps of the ATPase pathway. We used electron tomography of actively contracting insect flight muscle fast-frozen, freeze substituted, Araldite embedded, thin-sectioned and stained, to obtain 3D snapshots of the multiplicity of actin-attached myosin structures. We describe procedures for alignment of the repeating lattice of sub-volumes (38.7 nm cross-bridge repeats bounded by troponin) and multivariate data analysis to identify self-similar repeats for computing class averages. Improvements in alignment and classification of repeat sub-volumes reveals (for the first time in active muscle images) the helix of actin subunits in the thin filament and the troponin density with sufficient clarity that a quasiatomic model of the thin filament can be built into the class averages independent of the myosin cross-bridges. We show how quasiatomic model building can identify both strong and weak myosin attachments to actin. We evaluate the accuracy of image classification to enumerate the different types of actin-myosin attachments. | |||||||||
History |
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Structure visualization
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 532.4 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 11.3 KB 11.3 KB | Display Display | ![]() |
Images | ![]() | 189 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 199 KB | Display | ![]() |
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Full document | ![]() | 198.1 KB | Display | |
Data in XML | ![]() | 4.8 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 2w49MPC ![]() 2w4aMPC ![]() 2w4tMPC ![]() 2w4gC |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | This 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. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 6.9 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 : Isometrically contracting asynchronous insect flight muscle
Entire | Name: Isometrically contracting asynchronous insect flight muscle |
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Components |
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-Supramolecule #1000: Isometrically contracting asynchronous insect flight muscle
Supramolecule | Name: Isometrically contracting asynchronous insect flight muscle type: sample / ID: 1000 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 Oligomeric state: tissue / Number unique components: 8 |
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-Supramolecule #1: myofibril
Supramolecule | Name: myofibril / type: organelle_or_cellular_component / ID: 1 / Name.synonym: muscle Details: The sample was enblock stained using uranyl acetate and tannic acid, and post stained with lead citrate and potassium permanganate Oligomeric state: muscle fibril / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: ![]() |
-Experimental details
-Structure determination
Method | negative staining, cryo EM |
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![]() | electron tomography |
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Sample preparation
Buffer | Details: 20 mM MOPS buffer, 5 mM NaN3, and MgCl2, ATP, CaCl2, and EGTA in varying millimolar concentrations |
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Staining | Type: NEGATIVE Details: Freeze 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 ...Details: Freeze 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. |
Vitrification | Cryogen name: HELIUM / Chamber temperature: 4.5 K / Instrument: HOMEMADE PLUNGER Details: Vitrification instrument: modified Heuser Cryopress freezing head Method: smash against a liquid helium cooled Au-coated Cu mirror |
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Electron microscopy
Microscope | FEI/PHILIPS CM300FEG/T |
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Image recording | Category: CCD / Film or detector model: TVIPS TEMCAM-F224 (2k x 2k) Details: Images recorded on a TVIPS Tem-Cam F224 2k x 2k CCD camera Bits/pixel: 16 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm |
Sample stage | Specimen holder: Gatan model 670 Ultrahigh tilt analytical holder Specimen holder model: OTHER / Tilt series - Axis1 - Min angle: -72 ° / Tilt series - Axis1 - Max angle: 72 ° |
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Image processing
Final reconstruction | Algorithm: OTHER / Resolution method: FSC 0.5 CUT-OFF / Software - Name: 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 ...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. Number images used: 70 |
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