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- EMDB-14504: Three-dimensional structure of myosin binding protein C in rat ca... -

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Basic information

Entry
Database: EMDB / ID: EMD-14504
TitleThree-dimensional structure of myosin binding protein C in rat cardiac muscle
Map dataSubtomogram average of a 430 Angstrom repeat of C-zone
Sample
  • Tissue: Tokuyasu cryosection of cardiac muscle
Keywordsmuscle regulation / C-protiein / MyBP-C / hypertrophic cardiomyopathy / STRUCTURAL PROTEIN
Biological speciesRattus norvegicus (Norway rat)
Methodsubtomogram averaging / cryo EM / Resolution: 40.0 Å
AuthorsLuther PK / Morris EP / Huang X / Jun L
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
British Heart FoundationRG/11/21/29335 United Kingdom
CitationJournal: J Muscle Res Cell Motil / Year: 2023
Title: Cryo-electron tomography of intact cardiac muscle reveals myosin binding protein-C linking myosin and actin filaments.
Authors: Xinrui Huang / Iratxe Torre / Michele Chiappi / Zhan Yin / Anupama Vydyanath / Shuangyi Cao / Oliver Raschdorf / Morgan Beeby / Bonnie Quigley / Pieter P de Tombe / Jun Liu / Edward P Morris ...Authors: Xinrui Huang / Iratxe Torre / Michele Chiappi / Zhan Yin / Anupama Vydyanath / Shuangyi Cao / Oliver Raschdorf / Morgan Beeby / Bonnie Quigley / Pieter P de Tombe / Jun Liu / Edward P Morris / Pradeep K Luther /
Abstract: Myosin binding protein C (MyBP-C) is an accessory protein of the thick filament in vertebrate cardiac muscle arranged over 9 stripes of intervals of 430 Å in each half of the A-band in the region ...Myosin binding protein C (MyBP-C) is an accessory protein of the thick filament in vertebrate cardiac muscle arranged over 9 stripes of intervals of 430 Å in each half of the A-band in the region called the C-zone. Mutations in cardiac MyBP-C are a leading cause of hypertrophic cardiomyopathy the mechanism of which is unknown. It is a rod-shaped protein composed of 10 or 11 immunoglobulin- or fibronectin-like domains labelled C0 to C10 which binds to the thick filament via its C-terminal region. MyBP-C regulates contraction in a phosphorylation dependent fashion that may be through binding of its N-terminal domains with myosin or actin. Understanding the 3D organisation of MyBP-C in the sarcomere environment may provide new light on its function. We report here the fine structure of MyBP-C in relaxed rat cardiac muscle by cryo-electron tomography and subtomogram averaging of refrozen Tokuyasu cryosections. We find that on average MyBP-C connects via its distal end to actin across a disc perpendicular to the thick filament. The path of MyBP-C suggests that the central domains may interact with myosin heads. Surprisingly MyBP-C at Stripe 4 is different; it has weaker density than the other stripes which could result from a mainly axial or wavy path. Given that the same feature at Stripe 4 can also be found in several mammalian cardiac muscles and in some skeletal muscles, our finding may have broader implication and significance. In the D-zone, we show the first demonstration of myosin crowns arranged on a uniform 143 Å repeat.
History
DepositionMar 4, 2022-
Header (metadata) releaseMay 10, 2023-
Map releaseMay 10, 2023-
UpdateOct 11, 2023-
Current statusOct 11, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_14504.png

Downloads & links

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Map

FileDownload / File: emd_14504.map.gz / Format: CCP4 / Size: 6.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram average of a 430 Angstrom repeat of C-zone
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
5.14 Å/pix.
x 120 pix.
= 616.8 Å		5.14 Å/pix.
x 120 pix.
= 616.8 Å		5.14 Å/pix.
x 120 pix.
= 616.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 5.14 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.04
Minimum - Maximum-0.16164264 - 0.16977662
Average (Standard dev.)-0.00010810537 (±0.021307072)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions120120120
Spacing120120120
CellA=B=C: 616.8 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_14504_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_14504_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Tokuyasu cryosection of cardiac muscle

EntireName: Tokuyasu cryosection of cardiac muscle
Components
  • Tissue: Tokuyasu cryosection of cardiac muscle

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Supramolecule #1: Tokuyasu cryosection of cardiac muscle

SupramoleculeName: Tokuyasu cryosection of cardiac muscle / type: tissue / ID: 1 / Parent: 0 / Details: Longitudinal section of cardiac muscle
Source (natural)Organism: Rattus norvegicus (Norway rat) / Strain: Sprague Dawley / Organ: Heart / Tissue: Trabecula

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statetissue

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Sample preparation

BufferpH: 7.4
Component:
ConcentrationFormulaName
94.5 mmol/lNaClSodium chlorideSodium chloride
5.0 mmol/lKClPotassium chloride
25.0 mmol/lNaHCO3Sodium bicarbonate
1.0 mmol/lNa2HPO4Sodium phosphate
1.0 mmol/lMgSO4.7H20Magnesium sulphate
20.0 mmol/lNaCH3COOSodium acetate
1.0 mmol/lCaCl2Calcium chloride
30.0 mmol/l2,3-butanedione monoxime
5.0 mmol/lGlucose

Details: Krebs buffer was made fresh from concentrated components. It was aerated for 1/2 hour.
GridModel: Homemade / Material: NICKEL / Mesh: 200 / Support film - #0 - Film type ID: 1 / Support film - #0 - Material: CARBON / Support film - #0 - topology: CONTINUOUS / Support film - #1 - Film type ID: 2 / Support film - #1 - Material: FORMVAR / Support film - #1 - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.03 kPa
Details: The grid was coated with gold particles prior to freezing (recipe of Slot and Geuze).
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV
DetailsTrabeculae and papillary muscles were dissected from a rat heart

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 26000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 5.5 µm / Nominal defocus min: 4.0 µm / Nominal magnification: 26000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 88.0 K / Max: 88.0 K
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3840 pixel / Digitization - Dimensions - Height: 3712 pixel / Digitization - Frames/image: 1-12 / Number grids imaged: 1 / Average exposure time: 12.0 sec. / Average electron dose: 2.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

ExtractionNumber tomograms: 10 / Number images used: 1031 / Reference model: 1 selected subtomogram / Method: Volumes picked interactively at start / Software - Name: Dynamo
Final 3D classificationSoftware - Name: Dynamo
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionNumber classes used: 2 / Applied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 40.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: Dynamo / Number subtomograms used: 278
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: OTHER

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