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- EMDB-26462: cryo-EM structure of the rigor state wild type myosin-15-F-actin ... -

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Basic information

Entry
Database: EMDB / ID: EMD-26462
Titlecryo-EM structure of the rigor state wild type myosin-15-F-actin complex (symmetry expansion)
Map data
Sample
  • Complex: the rigor state wild type myosin-15-F-actin complex
    • Complex: Actin
    • Complex: Isoform 3 of Unconventional myosin-XV
    • Complex: chicken muscle regulatory light chain
Keywordsmyosin motor proteins / actin cytoskeleton / stereocilia / deafness / MOTOR PROTEIN
Biological speciesGallus gallus (chicken) / Mus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsGong R / Reynolds MJ / Alushin GM
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Dental and Craniofacial Research (NIH/NIDCR)DP5OD017885 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM141044 United States
National Institutes of Health/National Institute on Deafness and Other Communication Disorders (NIH/NIDCD)R01DC018827 United States
CitationJournal: Sci Adv / Year: 2022
Title: Structural basis for tunable control of actin dynamics by myosin-15 in mechanosensory stereocilia.
Authors: Rui Gong / Fangfang Jiang / Zane G Moreland / Matthew J Reynolds / Santiago Espinosa de Los Reyes / Pinar Gurel / Arik Shams / James B Heidings / Michael R Bowl / Jonathan E Bird / Gregory M Alushin /
Abstract: The motor protein myosin-15 is necessary for the development and maintenance of mechanosensory stereocilia, and mutations in myosin-15 cause hereditary deafness. In addition to transporting actin ...The motor protein myosin-15 is necessary for the development and maintenance of mechanosensory stereocilia, and mutations in myosin-15 cause hereditary deafness. In addition to transporting actin regulatory machinery to stereocilia tips, myosin-15 directly nucleates actin filament ("F-actin") assembly, which is disrupted by a progressive hearing loss mutation (p.D1647G, ""). Here, we present cryo-electron microscopy structures of myosin-15 bound to F-actin, providing a framework for interpreting the impacts of deafness mutations on motor activity and actin nucleation. Rigor myosin-15 evokes conformational changes in F-actin yet maintains flexibility in actin's D-loop, which mediates inter-subunit contacts, while the mutant locks the D-loop in a single conformation. Adenosine diphosphate-bound myosin-15 also locks the D-loop, which correspondingly blunts actin-polymerization stimulation. We propose myosin-15 enhances polymerization by bridging actin protomers, regulating nucleation efficiency by modulating actin's structural plasticity in a myosin nucleotide state-dependent manner. This tunable regulation of actin polymerization could be harnessed to precisely control stereocilium height.
History
DepositionMar 20, 2022-
Header (metadata) releaseAug 3, 2022-
Map releaseAug 3, 2022-
UpdateJan 17, 2024-
Current statusJan 17, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26462.png

Downloads & links

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Map

FileDownload / File: emd_26462.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.03 Å
Density
Contour LevelBy AUTHOR: 0.02
Minimum - Maximum-0.14698464 - 0.24217747
Average (Standard dev.)0.000005947933 (±0.00350054)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 527.36 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_26462_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_26462_half_map_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_26462_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : the rigor state wild type myosin-15-F-actin complex

EntireName: the rigor state wild type myosin-15-F-actin complex
Components
  • Complex: the rigor state wild type myosin-15-F-actin complex
    • Complex: Actin
    • Complex: Isoform 3 of Unconventional myosin-XV
    • Complex: chicken muscle regulatory light chain

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Supramolecule #1: the rigor state wild type myosin-15-F-actin complex

SupramoleculeName: the rigor state wild type myosin-15-F-actin complex / type: complex / ID: 1 / Parent: 0
Molecular weightTheoretical: 5.4 kDa/nm

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Supramolecule #2: Actin

SupramoleculeName: Actin / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: Gallus gallus (chicken)

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Supramolecule #3: Isoform 3 of Unconventional myosin-XV

SupramoleculeName: Isoform 3 of Unconventional myosin-XV / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Mus musculus (house mouse)

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Supramolecule #4: chicken muscle regulatory light chain

SupramoleculeName: chicken muscle regulatory light chain / type: complex / ID: 4 / Parent: 1
Source (natural)Organism: Gallus gallus (chicken)

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7
Details: 10 mM imidazole pH 7.0, 50 mM KCl, 1 mM MgCl2, 1 mM EGTA, 0.5 mM DTT, 0.01% NaN3
GridModel: C-flat-1.2/1.3 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: LEICA EM GP
Detailsthe rigor state wild type myosin-15 bound to F-actin

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 29000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-40 / Number grids imaged: 1 / Average exposure time: 10.0 sec. / Average electron dose: 60.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 427905
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6bno

Initial angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 3.0)
Final 3D classificationSoftware - Name: RELION (ver. 3.0)
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 3.0)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 223793
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6bno


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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