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- EMDB-7331: High-Resolution Cryo-EM Structures of Actin-bound Myosin States R... -

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Entry
Database: EMDB / ID: EMD-7331
TitleHigh-Resolution Cryo-EM Structures of Actin-bound Myosin States Reveal the Mechanism of Myosin Force Sensing
Map dataActin-bound Myosin
Sample
  • Complex: Complex of actin, myosin-1b, and calmodulin with ADP
    • Protein or peptide: Actin, alpha skeletal muscle
    • Protein or peptide: Unconventional myosin-Ib
    • Protein or peptide: Calmodulin
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
KeywordsMechanochemistry / Mechanobiology / Structural Biology / Cytoskeleton / Molecular Motor / Myosin-I / STRUCTURAL PROTEIN
Function / homology
Function and homology information


post-Golgi vesicle-mediated transport / transferrin transport / actin filament-based movement / vesicle transport along actin filament / CaM pathway / Cam-PDE 1 activation / myosin complex / Sodium/Calcium exchangers / Calmodulin induced events / Reduction of cytosolic Ca++ levels ...post-Golgi vesicle-mediated transport / transferrin transport / actin filament-based movement / vesicle transport along actin filament / CaM pathway / Cam-PDE 1 activation / myosin complex / Sodium/Calcium exchangers / Calmodulin induced events / Reduction of cytosolic Ca++ levels / Activation of Ca-permeable Kainate Receptor / CREB1 phosphorylation through the activation of CaMKII/CaMKK/CaMKIV cascasde / cytoskeletal motor activator activity / Loss of phosphorylation of MECP2 at T308 / microfilament motor activity / CREB1 phosphorylation through the activation of Adenylate Cyclase / PKA activation / negative regulation of high voltage-gated calcium channel activity / CaMK IV-mediated phosphorylation of CREB / Glycogen breakdown (glycogenolysis) / negative regulation of calcium ion export across plasma membrane / organelle localization by membrane tethering / Activation of RAC1 downstream of NMDARs / regulation of cardiac muscle cell action potential / mitochondrion-endoplasmic reticulum membrane tethering / CLEC7A (Dectin-1) induces NFAT activation / autophagosome membrane docking / tropomyosin binding / myosin heavy chain binding / mesenchyme migration / positive regulation of ryanodine-sensitive calcium-release channel activity / Negative regulation of NMDA receptor-mediated neuronal transmission / troponin I binding / regulation of cell communication by electrical coupling involved in cardiac conduction / Unblocking of NMDA receptors, glutamate binding and activation / negative regulation of peptidyl-threonine phosphorylation / Synthesis of IP3 and IP4 in the cytosol / actin filament bundle / Phase 0 - rapid depolarisation / filamentous actin / protein phosphatase activator activity / RHO GTPases activate PAKs / phosphatidylinositol-3,4,5-trisphosphate binding / actin filament bundle assembly / skeletal muscle thin filament assembly / microvillus / positive regulation of cyclic-nucleotide phosphodiesterase activity / brush border / cytoskeletal motor activity / striated muscle thin filament / positive regulation of phosphoprotein phosphatase activity / Long-term potentiation / Ion transport by P-type ATPases / Uptake and function of anthrax toxins / Calcineurin activates NFAT / Regulation of MECP2 expression and activity / skeletal muscle myofibril / actin monomer binding / catalytic complex / DARPP-32 events / detection of calcium ion / negative regulation of ryanodine-sensitive calcium-release channel activity / Smooth Muscle Contraction / RHO GTPases activate IQGAPs / regulation of cardiac muscle contraction / calcium channel inhibitor activity / cellular response to interferon-beta / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / Protein methylation / skeletal muscle fiber development / voltage-gated potassium channel complex / Activation of AMPK downstream of NMDARs / eNOS activation / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / stress fiber / regulation of calcium-mediated signaling / positive regulation of protein dephosphorylation / titin binding / regulation of ryanodine-sensitive calcium-release channel activity / Tetrahydrobiopterin (BH4) synthesis, recycling, salvage and regulation / Ion homeostasis / positive regulation of protein autophosphorylation / sperm midpiece / phosphatidylinositol-4,5-bisphosphate binding / calcium channel complex / substantia nigra development / actin filament polymerization / adenylate cyclase activator activity / Ras activation upon Ca2+ influx through NMDA receptor / regulation of heart rate / protein serine/threonine kinase activator activity / sarcomere / FCERI mediated Ca+2 mobilization / FCGR3A-mediated IL10 synthesis / trans-Golgi network membrane / VEGFR2 mediated vascular permeability / filopodium / positive regulation of peptidyl-threonine phosphorylation / Antigen activates B Cell Receptor (BCR) leading to generation of second messengers / VEGFR2 mediated cell proliferation
Similarity search - Function
Class I myosin tail homology domain / Class I myosin, motor domain / Unconventional myosin tail, actin- and lipid-binding / Class I myosin tail homology (TH1) domain profile. / IQ calmodulin-binding motif / 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. ...Class I myosin tail homology domain / Class I myosin, motor domain / Unconventional myosin tail, actin- and lipid-binding / Class I myosin tail homology (TH1) domain profile. / IQ calmodulin-binding motif / 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 / Actins signature 1. / Actin, conserved site / Actins signature 2. / Kinesin motor domain superfamily / Actin/actin-like conserved site / Actins and actin-related proteins signature. / Actin / Actin family / Actin / EF-hand domain pair / EF-hand, calcium binding motif / ATPase, nucleotide binding domain / 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
Calmodulin-1 / Actin, alpha skeletal muscle / Unconventional myosin-Ib
Similarity search - Component
Biological speciesunidentified (others) / Oryctolagus cuniculus (rabbit) / Rattus norvegicus (Norway rat)
Methodhelical reconstruction / cryo EM / Resolution: 3.9 Å
AuthorsMentes A / Huehn A
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-
UpdateMar 13, 2024-
Current statusMar 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.016
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.016
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6c1h
  • Surface level: 0.016
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6c1h
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_7331.png

Downloads & links

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Map

FileDownload / File: emd_7331.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationActin-bound Myosin
Voxel sizeX=Y=Z: 1.3 Å
Density
Contour LevelBy EMDB: 0.013 / Movie #1: 0.016
Minimum - Maximum-0.009556788 - 0.046719756
Average (Standard dev.)-0.00004045668 (±0.0022974052)
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-0.0100.047-0.000

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Supplemental data

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Mask #1

Fileemd_7331_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Actin-bound Myosin, half mask 1

Fileemd_7331_half_map_1.map
AnnotationActin-bound Myosin, half mask 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Actin-bound Myosin, half mask 2

Fileemd_7331_half_map_2.map
AnnotationActin-bound Myosin, half mask 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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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
Components
  • Complex: Complex of actin, myosin-1b, and calmodulin with ADP
    • Protein or peptide: Actin, alpha skeletal muscle
    • Protein or peptide: Unconventional myosin-Ib
    • Protein or peptide: Calmodulin
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION

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

SupramoleculeName: Complex of actin, myosin-1b, and calmodulin with ADP / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: unidentified (others)

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Macromolecule #1: Actin, alpha skeletal muscle

MacromoleculeName: Actin, alpha skeletal muscle / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Molecular weightTheoretical: 41.862613 KDa
SequenceString: DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IEHGIITNWD DMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSGDGV T HNVPIYEG ...String:
DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IEHGIITNWD DMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSGDGV T HNVPIYEG YALPHAIMRL DLAGRDLTDY LMKILTERGY SFVTTAEREI VRDIKEKLCY VALDFENEMA TAASSSSLEK SY ELPDGQV ITIGNERFRC PETLFQPSFI GMESAGIHET TYNSIMKCDI DIRKDLYANN VMSGGTTMYP GIADRMQKEI TAL APSTMK IKIIAPPERK YSVWIGGSIL ASLSTFQQMW ITKQEYDEAG PSIVHRKCF

UniProtKB: Actin, alpha skeletal muscle

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Macromolecule #2: Unconventional myosin-Ib

MacromoleculeName: Unconventional myosin-Ib / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 84.14393 KDa
SequenceString: VKSSLLDNMI GVGDTVLLEP LNEETFIDNL KKRFDHNEIY TYIGSVVISV NPYRSLPIYS PEKVEDYRNR NFYELSPHIF ALSDEAYRS LRDQDKDQCI LITGESGAGK TEASKLVMSY VAAVCGKGAE VNQVKEQLLQ STPVLEAFGN AKTVRNDNSS R FGKYMDIE ...String:
VKSSLLDNMI GVGDTVLLEP LNEETFIDNL KKRFDHNEIY TYIGSVVISV NPYRSLPIYS PEKVEDYRNR NFYELSPHIF ALSDEAYRS LRDQDKDQCI LITGESGAGK TEASKLVMSY VAAVCGKGAE VNQVKEQLLQ STPVLEAFGN AKTVRNDNSS R FGKYMDIE FDFKGDPLGG VISNYLLEKS RVVKQPRGER NFHVFYQLLS GASEELLHKL KLERDFSRYN YLSLDSAKVN GV DDAANFR TVRNAMQIVG FSDPEAESVL EVVAAVLKLG NIEFKPESRM NGLDESKIKD KNELKEICEL TSIDQVVLER AFS FRTVEA KQEKVSTTLN VAQAYYARDA LAKNLYSRLF SWLVNRINES IKAQTKVRKK VMGVLDIYGF EIFEDNSFEQ FIIN YCNEK LQQIFIELTL KEEQEEYIRE DIEWTHIDYF NNAIICDLIE NNTNGILAML DEECLRPGTV TDETFLEKLN QVCAT HQHF ESRMSKCSRF LNDTTLPHSC FRIQHYAGKV LYQVEGFVDK NNDLLYRDLS QAMWKAGHAL IKSLFPEGNP AKVNLK RPP TAGSQFKASV ATLMKNLQTK NPNYIRCIKP NDKKAAHIFS ESLVCHQIRY LGLLENVRVR RAGYAFRQAY EPCLERY KM LCKQTWPHWK GPARSGVEVL FNELEIPVEE YSFGRSKIFI RNPRTLFQLE DLRKQRLEDL ATLIQKIYRG WKCRTHFL L MKGLNDIF

UniProtKB: Unconventional myosin-Ib

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Macromolecule #3: Calmodulin

MacromoleculeName: Calmodulin / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: unidentified (others)
Molecular weightTheoretical: 16.72135 KDa
SequenceString:
ADQLTEEQIA EFKEAFSLFD KDGDGTITTK ELGTVMRSLG QNPTEAELQD MINEVDADGN GTIDFPEFLT MMARKMKDTD SEEEIREAF RVFDKDGNGY ISAAELRHVM TNLGEKLTDE EVDEMIREAD IDGDGQVNYE EFVQMMTAK

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Macromolecule #4: ADENOSINE-5'-DIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 5 / Formula: ADP
Molecular weightTheoretical: 427.201 Da
Chemical component information

ChemComp-ADP:
ADENOSINE-5'-DIPHOSPHATE / ADP, energy-carrying molecule*YM / Adenosine diphosphate

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Macromolecule #5: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 5 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statehelical array

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Sample preparation

BufferpH: 7
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average exposure time: 11.0 sec. / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final angle assignmentType: NOT APPLICABLE
Final reconstructionApplied symmetry - Helical parameters - Δz: 27.5 Å
Applied symmetry - Helical parameters - Δ&Phi: -167.4 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 3.9 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.
Number images used: 62000

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-6c1h:
High-Resolution Cryo-EM Structures of Actin-bound Myosin States Reveal the Mechanism of Myosin Force Sensing

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