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- PDB-6xe9: 10S myosin II (smooth muscle) -

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Basic information

Entry
Database: PDB / ID: 6xe9
Title10S myosin II (smooth muscle)
Components
  • Myosin II heavy chain (smooth muscle)
  • Myosin light chain 9
  • Myosin light chain smooth muscle isoform
KeywordsCONTRACTILE PROTEIN / motor protein / auto-inhibited form / 10 S conformation
Function / homology
Function and homology information


myosin complex / cytoskeletal motor activity / actin filament binding / calcium ion binding / ATP binding / cytosol
Similarity search - Function
EF-hand domain / DNA repair protein XRCC4-like, C-terminal / Myosin tail / Myosin tail / Myosin N-terminal SH3-like domain / Myosin S1 fragment, N-terminal / EF-hand domain pair / Myosin, N-terminal, SH3-like / Myosin N-terminal SH3-like domain profile. / Short calmodulin-binding motif containing conserved Ile and Gln residues. ...EF-hand domain / DNA repair protein XRCC4-like, C-terminal / Myosin tail / Myosin tail / Myosin N-terminal SH3-like domain / Myosin S1 fragment, N-terminal / EF-hand domain pair / Myosin, N-terminal, SH3-like / Myosin N-terminal SH3-like domain profile. / Short calmodulin-binding motif containing conserved Ile and Gln residues. / Myosin head, motor domain / Myosin head (motor domain) / Myosin motor domain profile. / Myosin. Large ATPases. / IQ motif profile. / IQ motif, EF-hand binding site / Kinesin motor domain superfamily / EF-hand domain pair / EF-hand, calcium binding motif / EF-Hand 1, calcium-binding site / EF-hand calcium-binding domain. / EF-hand calcium-binding domain profile. / EF-hand domain / EF-hand domain pair / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
MYH11 protein / EF-hand domain-containing protein / Myosin light chain smooth muscle isoform
Similarity search - Component
Biological speciesMeleagris gallopavo (turkey)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.3 Å
AuthorsTiwari, P. / Craig, R. / Padron, R.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS)R01AR072036 United States
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS)R01AR067279 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R01HL139883 United States
CitationJournal: Nature / Year: 2020
Title: Cryo-EM structure of the inhibited (10S) form of myosin II.
Authors: Shixin Yang / Prince Tiwari / Kyoung Hwan Lee / Osamu Sato / Mitsuo Ikebe / Raúl Padrón / Roger Craig /
Abstract: Myosin II is the motor protein that enables muscle cells to contract and nonmuscle cells to move and change shape. The molecule has two identical heads attached to an elongated tail, and can exist in ...Myosin II is the motor protein that enables muscle cells to contract and nonmuscle cells to move and change shape. The molecule has two identical heads attached to an elongated tail, and can exist in two conformations: 10S and 6S, named for their sedimentation coefficients. The 6S conformation has an extended tail and assembles into polymeric filaments, which pull on actin filaments to generate force and motion. In 10S myosin, the tail is folded into three segments and the heads bend back and interact with each other and the tail, creating a compact conformation in which ATPase activity, actin activation and filament assembly are all highly inhibited. This switched-off structure appears to function as a key energy-conserving storage molecule in muscle and nonmuscle cells, which can be activated to form functional filaments as needed-but the mechanism of its inhibition is not understood. Here we have solved the structure of smooth muscle 10S myosin by cryo-electron microscopy with sufficient resolution to enable improved understanding of the function of the head and tail regions of the molecule and of the key intramolecular contacts that cause inhibition. Our results suggest an atomic model for the off state of myosin II, for its activation and unfolding by phosphorylation, and for understanding the clustering of disease-causing mutations near sites of intramolecular interaction.
History
DepositionJun 12, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 2, 2020Provider: repository / Type: Initial release
Revision 1.1Dec 16, 2020Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed ..._citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Dec 30, 2020Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.3Mar 6, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

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  • Deposited structure unit
  • Imaged by Jmol
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  • Superimposition on EM map
  • EMDB-22145
  • Imaged by UCSF Chimera
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Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Myosin II heavy chain (smooth muscle)
B: Myosin light chain smooth muscle isoform
C: Myosin light chain 9
M: Myosin II heavy chain (smooth muscle)
N: Myosin light chain smooth muscle isoform
O: Myosin light chain 9


Theoretical massNumber of molelcules
Total (without water)532,0196
Polymers532,0196
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: Native molecular weight combined with chain weights. Single molecule EM of intact molecule.
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area28340 Å2
ΔGint-234 kcal/mol
Surface area144420 Å2

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Components

#1: Antibody Myosin II heavy chain (smooth muscle)


Mass: 229148.250 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Meleagris gallopavo (turkey) / References: UniProt: G1N5L2*PLUS, myosin ATPase
#2: Protein Myosin light chain smooth muscle isoform


Mass: 16989.145 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Meleagris gallopavo (turkey) / References: UniProt: Q6W5H0
#3: Protein Myosin light chain 9 /


Mass: 19872.244 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Meleagris gallopavo (turkey) / References: UniProt: G3URE9

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Smooth muscle myosin II / Type: COMPLEX / Details: Whole myosin II isolated from turkey gizzard / Entity ID: all / Source: NATURAL
Molecular weightValue: 0.53 MDa / Experimental value: NO
Source (natural)Organism: Meleagris gallopavo (turkey) / Organ: gizzard / Tissue: smooth muscle
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
10.15 Msodium acetateCH3COONa1
21 mMEGTAC14H24N2O101
32.5 mMmagnesium chlorideMgCl21
40.5 mMadenosine triphosphateC10H16N5O13P31
510 mMMOPSC7H15NO4S1
SpecimenConc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: The specimen was cross-linked in presence of 0.1% glutaraldehyde for 60 seconds.
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 283.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 105000 X / Nominal defocus max: 3500 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 43.3 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 10951
Image scansWidth: 5760 / Height: 4092

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.17.1_3660refinement
PHENIX1.17.1_3660refinement
EM software
IDNameVersionCategoryDetails
1RELION3particle selection
2SerialEM3.8.0image acquisition
4CTFFIND4.1.13CTF correction
7UCSF Chimera1.14model fittingFit model in map
9RELION3initial Euler assignment
10RELION3final Euler assignment
11RELION3classification
12RELION33D reconstruction
19Coot0.8.9.2 ELmodel refinementCoot was used for regularization and local refinement
20PHENIX1.17.1 - 3660model refinementIt was used for Real space refinement and model validation
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1765220
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 168613 / Algorithm: BACK PROJECTION / Num. of class averages: 3 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
11I84

1i84

11I841PDBexperimental model
22FXM

2fxm

12FXM2PDBexperimental model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 257.18 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.002522413
ELECTRON MICROSCOPYf_angle_d0.47530291
ELECTRON MICROSCOPYf_chiral_restr0.03633419
ELECTRON MICROSCOPYf_plane_restr0.00274025
ELECTRON MICROSCOPYf_dihedral_angle_d6.19833127

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