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Molecular Models of Averaged Rigor Crossbridges from Tomograms of Insect Flight Muscle

by electron tomography, at 70 A resolution

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Entry
Summary
Database / IDPORTEIN DATA BANK (PDB) / 1m8q
TitleMolecular Models of Averaged Rigor Crossbridges from Tomograms of Insect Flight Muscle
DescriptorChicken Skeletal muscle Myosin II
Chicken Skeletal muscle Myosin II Regulatory Light Chain
Chicken Skeletal muscle Myosin II Essential Light Chain
Rabbit Skeletal muscle Actin
KeywordsCONTRACTILE PROTEIN, Actin-myosin complex in situ in muscle
AuthorsChen, L.F., Winkler, H., Reedy, M.K., Reedy, M.C., Taylor, K.A.
DateDeposition: 2002-07-25, Release: 2002-09-10
PDBj Mine pagesSummary, Structural Details, Experimental Details, Functional Details
Other databasesRCSB-PDB, PDBe, CATH, CE, FSSP, SCOP, VAST
Structure Visualization
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List of similar structure data about Omokage system
Article
Citation - primary
ArticleJ. Struct. Biol., Vol. 138, Issue 1-2, Page 92-104
TitleMolecular modeling of averaged rigor crossbridges from tomograms of insect flight muscle.
AuthorsLi Fan Chen, Hanspeter Winkler, Michael K Reedy, Mary C Reedy, Kenneth A Taylor
Institute of Molecular Biophysics, Florida State University, Tallahassee, FL 32306-4380, USA.
KeywordsActomyosin (chemistry, 9013-26-7), Animals, Flight, Animal, Hemiptera, Insects, Microscopy, Electron, Models, Molecular, Muscle Fibers, Skeletal (chemistry), Muscle Proteins (chemistry), Muscle, Skeletal (chemistry), Tomography, X-Ray Computed
LinksPubMed: 12160705, PII: S1047847702000138
Citation - 1
ArticleJ.STRUCT.BIOL., Vol. 133, Page: 221 - 232, Year: 2001,
TitleReal space refinement of acto-myosin structures from sectioned muscle.
AuthorsChen, L.F., Blanc, E., Chapman, M.S., Taylor, K.A.
LinksDOI: 10.1006/jsbi.2000.4321
Citation - 2
ArticleULTRAMICROSCOPY, Vol. 77, Page: 141 - 152, Year: 1999,
TitleMultivariate statistical analysis of three-dimensional cross-bridge motifs in insect flight muscle
AuthorsWinkler, H., Taylor, K.A.
LinksDOI: 10.1016/S0304-3991(99)00035-2
Citation - 3
ArticleJ.STRUCT.BIOL., Vol. 120, Page: 372 - 386, Year: 1997,
TitleThe use of electron tomography for structural analysis of disordered protein arrays.
AuthorsTaylor, K.A., Tang, J., Cheng, Y., Winkler, H.
LinksDOI: 10.1006/jsbi.1997.3932
Components
ID 1 : Skeletal muscle Myosin II
Image
DescriptionSkeletal muscle Myosin II
Typepolypeptide(L)
Formula weight96626.203 Da
Number of molecules4
ID1
SourceMethod: Isolated from a natural source
Common name: chicken
Details: Organism from which the myosin for the crystal structure that provided 2MYS was obtained
Genus: Gallus
NCBI taxonomy: ID:9031
Organism scientific: Gallus gallus

LinksUniProt: P13538, Sequence view
ID 2 : Skeletal muscle Myosin II Regulatory Light Chain
Image
DescriptionSkeletal muscle Myosin II Regulatory Light Chain
Typepolypeptide(L)
Formula weight16063.167 Da
Number of molecules4
ID2
SourceMethod: Isolated from a natural source
Common name: chicken
Details: Organism from which the myosin for the crystal structure that provided 2MYS was obtained
Genus: Gallus
NCBI taxonomy: ID:9031
Organism scientific: Gallus gallus
LinksUniProt: P02609, Sequence view
ID 3 : Skeletal muscle Myosin II Essential Light Chain
Image
DescriptionSkeletal muscle Myosin II Essential Light Chain
Typepolypeptide(L)
Formula weight16064.055 Da
Number of molecules4
ID3
SourceMethod: Isolated from a natural source
Common name: chicken
Details: Organism from which the Myosin for the crystal structure that provided 2MYS was obtained
Genus: Gallus
NCBI taxonomy: ID:9031
Organism scientific: Gallus gallus
LinksUniProt: P02604, Sequence view
ID 4 : Skeletal muscle Actin
Image
DescriptionSkeletal muscle Actin
Typepolypeptide(L)
Formula weight41862.965 Da
Number of molecules14
ID4
SourceMethod: Isolated from a natural source
Common name: rabbit
Details: Organism from which the Actin that provided the crystal structure for 1ATN was obtained
Genus: Oryctolagus
NCBI taxonomy: ID:9986
Organism scientific: Oryctolagus cuniculus

LinksUniProt: P68135, Sequence view
Sample
Assembly
Aggregation stateTISSUE
DetailsDorsal longitudinal flight muscles of the large waterbug, Lethocerus maximus were glycerinated in the dissected thorax and stored at -20 degrees C. Fixation involved sequential tannic acid-glutaraldehyde (0.2% and 2%), cold 1% OsO4 at pH 6.0, 1% uranyl acetate block staining and Araldite 506 emgedding. Thin sections were stained with a sequence of permanganate and lead. Section were ~25 nm thick.
NameInsect flight muscle
Entity assembly
Assembly-ID1
DetailsModels are built to fit the thin, actin containing filaments labeled with endogenous myosin in the rigor state
NameInsect flight muscle of the large water bug Lethocerus maximus
Experiment
Reconstruction methodTOMOGRAPHY
Specimen typeFREEZE SUBSTITUTION
Vitrification
DetailsNo vitrification. Samples were viewed at room temperature.
Experiment
MethodELECTRON MICROSCOPY
Electron Microscopy
Detector
TypeKodak S0163 Film
Image scans
Number digital images36
Processing
2D projection selection
Software nameRSref
3D reconstruction
CTF correction methodnone
Details3-D motifs were identified in the tomogram by first producing a cross correlation map from which peak coordinates were determined from their center of gravity. We define a 3-D motif as one, entire 38.7 nm crossbridge repeat along actin. These motifs usually contain at least four myosin heads in two paired crossbridges (single chevrons) and sometimes contain as many as six myosin heads in four paired crossbridges (double chevrons). The reference for the analysis was selected to be centered between successive troponin densities which could be identified from the in-plane projection. The individual crossbridge motifs were then subjected to multivariate statistical analysis to identify clusters of motifs showing similar crossbridge structure. These clusters formed the class averages. The choice of structure to be classified was decided by the resolution and the later process of model building. Averaging was done according to the heirarchical ascendent method. The resolution in each of the class averages was 7 nm by the spectral signal to noise ratio.
Magnification calibrationIndicated instrument magnification
MethodDual axis tilt series electron tomography
Nominal pixel size15.5 A/pix
Resolution70 A
3D fitting
MethodInitial models were fit by hand using "O". The fit was then refined using real space refinement.
Refinement Protocolrigid body
Refinement SpaceREAL
Software nameRSref
Target criteriaBest correlation coefficient and fewest poor contacts
3D fitting list
3D Fitting ID1
Refine hist
Cycle idLAST
Refine idELECTRON MICROSCOPY
Total atoms76872
Protein atoms76872
Download
PDB format
Allpdb1m8q.ent.gz
pdb1m8q.ent (uncompressed file)
Header onlypdb1m8q.ent.gz
mmCIF format
mmCIF1m8q.cif.gz
XML format
All1m8q.xml.gz
No-atom1m8q-noatom.xml.gz
Ext-atom1m8q-extatom.xml.gz
Movie files
movie #1
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