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- EMDB-16848: Cryo-EM structure of actomyosin-5a-S1 with the full-length lever ... -

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Basic information

Entry
Database: EMDB / ID: EMD-16848
TitleCryo-EM structure of actomyosin-5a-S1 with the full-length lever (nucleotide free, class B)
Map dataDeepEMhancer post-processed map
Sample
  • Complex: Myosin-5a bound to F-actin
    • Complex: Actin, alpha skeletal muscle
    • Complex: Unconventional myosin-Va with calmodulin-1 bound
      • Complex: Calmodulin-1
Keywordsmyosin / cytoskeletal motor / myosin-va / myo5a / myosin-5a / S1 / rigor / nucleotide free / apo / lever / 6IQ / actomyosin / actomyosin-5a / actin / actomyosin-va / actin bound / MOTOR PROTEIN
Biological speciesOryctolagus cuniculus (rabbit) / Mus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 8.1 Å
AuthorsGravett MSC / Klebl DP / Harlen OG / Read DJ / Harris SA / Muench SP / Peckham M
Funding support United Kingdom, 5 items
OrganizationGrant numberCountry
Wellcome Trust102174/B/13/Z United Kingdom
Wellcome Trust223125/Z/21/Z United Kingdom
Wellcome Trust108466/Z/15/Z United Kingdom
Engineering and Physical Sciences Research CouncilEP/T022167/1 United Kingdom
Engineering and Physical Sciences Research CouncilEP/R013012 United Kingdom
CitationJournal: Structure / Year: 2024
Title: Exploiting cryo-EM structures of actomyosin-5a to reveal the physical properties of its lever.
Authors: Molly S C Gravett / David P Klebl / Oliver G Harlen / Daniel J Read / Stephen P Muench / Sarah A Harris / Michelle Peckham /
Abstract: Myosin 5a (Myo5a) is a dimeric processive motor protein that transports cellular cargos along filamentous actin (F-actin). Its long lever is responsible for its large power-stroke, step size, and ...Myosin 5a (Myo5a) is a dimeric processive motor protein that transports cellular cargos along filamentous actin (F-actin). Its long lever is responsible for its large power-stroke, step size, and load-bearing ability. Little is known about the levers' structure and physical properties, and how they contribute to walking mechanics. Using cryoelectron microscopy (cryo-EM) and molecular dynamics (MD) simulations, we resolved the structure of monomeric Myo5a, comprising the motor domain and full-length lever, bound to F-actin. The range of its lever conformations revealed its physical properties, how stiffness varies along its length and predicts a large, 35 nm, working stroke. Thus, the newly released trail head in a dimeric Myo5a would only need to perform a small diffusive search for its new binding site on F-actin, and stress would only be generated across the dimer once phosphate is released from the lead head, revealing new insight into the walking behavior of Myo5a.
History
DepositionMar 14, 2023-
Header (metadata) releaseSep 25, 2024-
Map releaseSep 25, 2024-
UpdateApr 9, 2025-
Current statusApr 9, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16848.png

Downloads & links

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Map

FileDownload / File: emd_16848.map.gz / Format: CCP4 / Size: 59.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDeepEMhancer post-processed map
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
2.13 Å/pix.
x 250 pix.
= 532.5 Å		2.13 Å/pix.
x 250 pix.
= 532.5 Å		2.13 Å/pix.
x 250 pix.
= 532.5 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 2.13 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.28
Minimum - Maximum-0.0005694516 - 1.4949923
Average (Standard dev.)0.0013553416 (±0.024763798)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions250250250
Spacing250250250
CellA=B=C: 532.5 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_16848_msk_1.map
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Additional map: Masked post-processed map

Fileemd_16848_additional_1.map
AnnotationMasked post-processed map
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Additional map: Gaussian filtered (SD 5) map

Fileemd_16848_additional_2.map
AnnotationGaussian filtered (SD 5) map
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Additional map: Raw map

Fileemd_16848_additional_3.map
AnnotationRaw map
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Half map: Half map 2

Fileemd_16848_half_map_1.map
AnnotationHalf map 2
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Half map: Half map 1

Fileemd_16848_half_map_2.map
AnnotationHalf map 1
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Sample components

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Entire : Myosin-5a bound to F-actin

EntireName: Myosin-5a bound to F-actin
Components
  • Complex: Myosin-5a bound to F-actin
    • Complex: Actin, alpha skeletal muscle
    • Complex: Unconventional myosin-Va with calmodulin-1 bound
      • Complex: Calmodulin-1

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Supramolecule #1: Myosin-5a bound to F-actin

SupramoleculeName: Myosin-5a bound to F-actin / type: complex / ID: 1 / Parent: 0
Molecular weightTheoretical: 20 KDa

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Supramolecule #2: Actin, alpha skeletal muscle

SupramoleculeName: Actin, alpha skeletal muscle / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: Oryctolagus cuniculus (rabbit) / Organ: skeletal muscle

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Supramolecule #3: Unconventional myosin-Va with calmodulin-1 bound

SupramoleculeName: Unconventional myosin-Va with calmodulin-1 bound / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Mus musculus (house mouse)

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Supramolecule #4: Calmodulin-1

SupramoleculeName: Calmodulin-1 / type: complex / ID: 4 / Parent: 3 / Details: 1 calmodulin bound per IQ domain (6 overall)

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statehelical array

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Sample preparation

BufferpH: 7
Component:
ConcentrationFormulaName
100.0 mMKClpotassium chloride
0.1 mMC14H24N2O10egtazic acid (EGTA)
1.0 mMMgCl2magnesium chloride
10.0 mMC7H15NO4S3-morpholinopropane-1-sulfonic acid (MOPS)
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AMYLAMINE
VitrificationCryogen name: ETHANE / Chamber humidity: 80 % / Chamber temperature: 281.15 K / Instrument: FEI VITROBOT MARK IV
Details5.26 mg/mL Actin, alpha skeletal muscle 266 mg/mL FlAG-tagged unconventional myosin-Va (residues 1-907) 84.2 mg/mL Calmodulin-1

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: INTEGRATING / Number grids imaged: 1 / Number real images: 4285 / Average exposure time: 1.5 sec. / Average electron dose: 63.13 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.6 µm / Nominal defocus min: 1.8 µm / Nominal magnification: 75000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 838213
Startup modelType of model: OTHER / Details: relion_reconstruct map from particles in 3D class
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 8.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 18695
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 3D classificationNumber classes: 20 / Software - Name: RELION (ver. 3.1)
FSC plot (resolution estimation)

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