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- EMDB-7330: High-Resolution Cryo-EM Structures of Actin-bound Myosin States R... -

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Basic information

Entry
Database: EMDB / ID: EMD-7330
TitleHigh-Resolution Cryo-EM Structures of Actin-bound Myosin States Reveal the Mechanism of Myosin Force Sensing
Map data
SampleComplex of actin, myosin-1b, and calmodulin with ADP:
Unconventional myosin-Ib / Calmodulin / Actin, alpha skeletal muscle / (ligand) x 2
Function / homology
Function and homology information


actin filament-based movement / transferrin transport / vesicle transport along actin filament / CaM pathway / Sodium/Calcium exchangers / Cam-PDE 1 activation / Reduction of cytosolic Ca++ levels / microfilament motor activity / Calmodulin induced events / => GO:0120081 ...actin filament-based movement / transferrin transport / vesicle transport along actin filament / CaM pathway / Sodium/Calcium exchangers / Cam-PDE 1 activation / Reduction of cytosolic Ca++ levels / microfilament motor activity / Calmodulin induced events / => GO:0120081 / microfilament motor activity => GO:0000146 / Glycogen breakdown (glycogenolysis) / CREB1 phosphorylation through the activation of CaMKII/CaMKK/CaMKIV cascasde / myosin complex / CLEC7A (Dectin-1) induces NFAT activation / Activation of Ca-permeable Kainate Receptor / Loss of phosphorylation of MECP2 at T308 / glycogen catabolic process / PKA activation / CREB1 phosphorylation through the activation of Adenylate Cyclase / mesenchyme migration / establishment of protein localization to mitochondrial membrane / organelle localization by membrane tethering / CaMK IV-mediated phosphorylation of CREB / regulation of cardiac muscle cell action potential / negative regulation of high voltage-gated calcium channel activity / N-terminal myristoylation domain binding / autophagosome membrane docking / tropomyosin binding / mitochondrion-endoplasmic reticulum membrane tethering / myosin heavy chain binding / Phase 0 - rapid depolarisation / Activation of RAC1 downstream of NMDARs / troponin I binding / type 3 metabotropic glutamate receptor binding / regulation of cell communication by electrical coupling involved in cardiac conduction / Ion transport by P-type ATPases / regulation of synaptic vesicle endocytosis / Negative regulation of NMDA receptor-mediated neuronal transmission / negative regulation of peptidyl-threonine phosphorylation / Unblocking of NMDA receptors, glutamate binding and activation / protein phosphatase activator activity / brush border / Synthesis of IP3 and IP4 in the cytosol / positive regulation of ryanodine-sensitive calcium-release channel activity / skeletal muscle thin filament assembly / actin filament bundle / striated muscle thin filament / filamentous actin / regulation of rhodopsin mediated signaling pathway / Smooth Muscle Contraction / response to corticosterone / catalytic complex / RHO GTPases activate PAKs / positive regulation of cyclic-nucleotide phosphodiesterase activity / Activation of AMPK downstream of NMDARs / adenylate cyclase binding / Long-term potentiation / inositol phosphate metabolic process / detection of calcium ion / phosphatidylinositol-3,4,5-trisphosphate binding / actin filament bundle assembly / actin monomer binding / Uptake and function of anthrax toxins / Calcineurin activates NFAT / Regulation of MECP2 expression and activity / Protein methylation / cytoskeletal motor activity / skeletal muscle fiber development / negative regulation of ryanodine-sensitive calcium-release channel activity / regulation of cardiac muscle contraction / DARPP-32 events / skeletal muscle myofibril / adenylate cyclase activator activity / RHO GTPases activate IQGAPs / post-Golgi vesicle-mediated transport / calcium channel inhibitor activity / nitric-oxide synthase binding / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / positive regulation of phosphoprotein phosphatase activity / activation of adenylate cyclase activity / Inactivation, recovery and regulation of the phototransduction cascade / Ion homeostasis / stress fiber / trans-Golgi network membrane / voltage-gated potassium channel complex / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / regulation of heart rate / Wnt signaling pathway, calcium modulating pathway / titin binding / tetrahydrobiopterin metabolic process / substantia nigra development / eNOS activation / sarcomere / actin filament polymerization / positive regulation of protein dephosphorylation / calcium channel complex / Tetrahydrobiopterin (BH4) synthesis, recycling, salvage and regulation / phosphatidylinositol 3-kinase binding / regulation of ryanodine-sensitive calcium-release channel activity
Similarity search - Function
Class I myosin tail homology (TH1) domain profile. / Unconventional myosin tail, actin- and lipid-binding / Class I myosin tail homology domain / Class I myosin, motor domain / IQ calmodulin-binding motif / Short calmodulin-binding motif containing conserved Ile and Gln residues. / Myosin. Large ATPases. / Myosin head, motor domain / Myosin head (motor domain) / Myosin motor domain profile. ...Class I myosin tail homology (TH1) domain profile. / Unconventional myosin tail, actin- and lipid-binding / Class I myosin tail homology domain / Class I myosin, motor domain / IQ calmodulin-binding motif / Short calmodulin-binding motif containing conserved Ile and Gln residues. / Myosin. Large ATPases. / Myosin head, motor domain / Myosin head (motor domain) / Myosin motor domain profile. / IQ motif profile. / IQ motif, EF-hand binding site / Actins signature 1. / Actin, conserved site / Actins signature 2. / Actins and actin-related proteins signature. / Actin/actin-like conserved site / Actin / Actin / Actin family / Kinesin motor domain superfamily / EF-hand domain pair / ATPase, nucleotide binding domain / EF-hand, calcium binding motif / EF-Hand 1, calcium-binding site / EF-hand calcium-binding domain profile. / EF-hand domain / EF-hand calcium-binding domain. / EF-hand domain pair / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Calmodulin-1 / Actin, alpha skeletal muscle / Unconventional myosin-Ib
Similarity search - Component
Biological speciesunidentified (others) / Rat (rat) / Rabbit (rabbit)
Methodhelical reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsMentes A / Huehn A / Liu X / Zwolak A / Dominguez R / Shuman H / Ostap EM / Sindelar CV
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R37 GM057247 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2018
Title: High-resolution cryo-EM structures of actin-bound myosin states reveal the mechanism of myosin force sensing.
Authors: Ahmet Mentes / Andrew Huehn / Xueqi Liu / Adam Zwolak / Roberto Dominguez / Henry Shuman / E Michael Ostap / Charles V Sindelar /
Abstract: Myosins adjust their power outputs in response to mechanical loads in an isoform-dependent manner, resulting in their ability to dynamically adapt to a range of motile challenges. Here, we reveal the ...Myosins adjust their power outputs in response to mechanical loads in an isoform-dependent manner, resulting in their ability to dynamically adapt to a range of motile challenges. Here, we reveal the structural basis for force-sensing based on near-atomic resolution structures of one rigor and two ADP-bound states of myosin-IB (myo1b) bound to actin, determined by cryo-electron microscopy. The two ADP-bound states are separated by a 25° rotation of the lever. The lever of the first ADP state is rotated toward the pointed end of the actin filament and forms a previously unidentified interface with the N-terminal subdomain, which constitutes the upper half of the nucleotide-binding cleft. This pointed-end orientation of the lever blocks ADP release by preventing the N-terminal subdomain from the pivoting required to open the nucleotide binding site, thus revealing how myo1b is inhibited by mechanical loads that restrain lever rotation. The lever of the second ADP state adopts a rigor-like orientation, stabilized by class-specific elements of myo1b. We identify a role for this conformation as an intermediate in the ADP release pathway. Moreover, comparison of our structures with other myosins reveals structural diversity in the actomyosin binding site, and we reveal the high-resolution structure of actin-bound phalloidin, a potent stabilizer of filamentous actin. These results provide a framework to understand the spectrum of force-sensing capacities among the myosin superfamily.
History
DepositionJan 4, 2018-
Header (metadata) releaseJan 31, 2018-
Map releaseJan 31, 2018-
UpdateJan 8, 2020-
Current statusJan 8, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.017
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.017
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6c1g
  • Surface level: 0.017
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6c1g
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_7330.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.3 Å/pix.
x 400 pix.
= 520. Å
1.3 Å/pix.
x 400 pix.
= 520. Å
1.3 Å/pix.
x 400 pix.
= 520. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.3 Å
Density
Contour LevelBy EMDB: 0.015 / Movie #1: 0.017
Minimum - Maximum-0.023726193 - 0.07133766
Average (Standard dev.)0.00008748186 (±0.0027548608)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 520.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.31.31.3
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z520.000520.000520.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.0240.0710.000

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Supplemental data

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Segmentation: #1

Fileemd_7330_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Actin-bound Myosin, half mask 1

Fileemd_7330_half_map_1.map
AnnotationActin-bound Myosin, half mask 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Actin-bound Myosin, half mask 2

Fileemd_7330_half_map_2.map
AnnotationActin-bound Myosin, half mask 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire Complex of actin, myosin-1b, and calmodulin with ADP

EntireName: Complex of actin, myosin-1b, and calmodulin with ADP / Number of Components: 6

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Component #1: protein, Complex of actin, myosin-1b, and calmodulin with ADP

ProteinName: Complex of actin, myosin-1b, and calmodulin with ADP / Recombinant expression: No
SourceSpecies: unidentified (others)

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Component #2: protein, Unconventional myosin-Ib

ProteinName: Unconventional myosin-Ib / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 84.14393 kDa
SourceSpecies: Rat (rat)

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Component #3: protein, Calmodulin

ProteinName: Calmodulin / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 16.72135 kDa
SourceSpecies: unidentified (others)

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Component #4: protein, Actin, alpha skeletal muscle

ProteinName: Actin, alpha skeletal muscle / Number of Copies: 5 / Recombinant expression: No
MassTheoretical: 41.862613 kDa
SourceSpecies: Rabbit (rabbit)

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Component #5: ligand, MAGNESIUM ION

LigandName: MAGNESIUM ION / Number of Copies: 6 / Recombinant expression: No
MassTheoretical: 2.430505 MDa

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Component #6: ligand, ADENOSINE-5'-DIPHOSPHATE

LigandName: ADENOSINE-5'-DIPHOSPHATE / Number of Copies: 6 / Recombinant expression: No
MassTheoretical: 0.427201 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen State: helical array / Method: cryo EM
Helical ParametersAxial Symmetry: C1 (asymmetric) / Delta Z: 27.5 Å / Delta Phi: -167.4 %deg;
Sample solutionpH: 7
VitrificationCryogen Name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron Source: FIELD EMISSION GUN / Accelerating Voltage: 300 kV / Electron Dose: 50 e/Å2 / Illumination Mode: SPOT SCAN
LensImaging Mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: helical reconstruction
3D reconstructionResolution: 3.8 Å / Resolution Method: FSC 0.143 CUT-OFF
Details: Resolution estimated by post-processing in RELION using a mask with soft edges that included only the central subunit.

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Atomic model buiding

Modeling #1Refinement protocol: flexible / Refinement space: REAL
Output model

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