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- EMDB-26007: Three-dimensional organization of the apical complex in Toxoplasm... -

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Entry
Database: EMDB / ID: EMD-26007
TitleThree-dimensional organization of the apical complex in Toxoplasma tachyzoites
Map data
Sample
  • Cell: 3D reconstruction and annotation of distinct secretory organelles and cytoskeleton elements in the apical region of Toxoplasma gondii
KeywordsParasites / Toxoplasma gondii / apical end / cryoET / CELL INVASION
Biological speciesToxoplasma gondii (eukaryote)
Methodelectron tomography / cryo EM
AuthorsSun SY / Segev-Zarko L
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)S10OD021600 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P41GM103832 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM079429 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P01GM121203 United States
National Institutes of Health/National Institute of Mental Health (NIH/NIMH)R21MH125285-01A1 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2022
Title: Cryo-ET of parasites gives subnanometer insight into tubulin-based structures.
Authors: Stella Y Sun / Li-Av Segev-Zarko / Muyuan Chen / Grigore D Pintilie / Michael F Schmid / Steven J Ludtke / John C Boothroyd / Wah Chiu /
Abstract: Tubulin is a conserved protein that polymerizes into different forms of filamentous structures in , an obligate intracellular parasite in the phylum Apicomplexa. Two key tubulin-containing ...Tubulin is a conserved protein that polymerizes into different forms of filamentous structures in , an obligate intracellular parasite in the phylum Apicomplexa. Two key tubulin-containing cytoskeletal components are subpellicular microtubules (SPMTs) and conoid fibrils (CFs). The SPMTs help maintain shape and gliding motility, while the CFs are implicated in invasion. Here, we use cryogenic electron tomography to determine the molecular structures of the SPMTs and CFs in vitrified intact and detergent-extracted parasites. Subvolume densities from detergent-extracted parasites yielded averaged density maps at subnanometer resolutions, and these were related back to their architecture in situ. An intralumenal spiral lines the interior of the 13-protofilament SPMTs, revealing a preferred orientation of these microtubules relative to the parasite's long axis. Each CF is composed of nine tubulin protofilaments that display a comma-shaped cross-section, plus additional associated components. Conoid protrusion, a crucial step in invasion, is associated with an altered pitch of each CF. The use of basic building blocks of protofilaments and different accessory proteins in one organism illustrates the versatility of tubulin to form two distinct types of assemblies, SPMTs and CFs.
History
DepositionJan 21, 2022-
Header (metadata) releaseMar 2, 2022-
Map releaseMar 2, 2022-
UpdateJan 17, 2024-
Current statusJan 17, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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Supplemental images

emd_26007.png

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Map

FileDownload / File: emd_26007.map.gz / Format: CCP4 / Size: 1 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
14.17 Å/pix.
x 305 pix.
= 4322.46 Å		14.17 Å/pix.
x 959 pix.
= 13590.948 Å		14.17 Å/pix.
x 927 pix.
= 13137.444 Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 14.172 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-3.0 - 3.0
Average (Standard dev.)-0.0004954568 (±0.9724208)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions959927305
Spacing927959305
CellA: 13137.444 Å / B: 13590.948 Å / C: 4322.46 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z14.17214.17214.172
M x/y/z927959305
origin x/y/z0.0000.0000.000
length x/y/z13137.44413590.9484322.460
α/β/γ90.00090.00090.000
start NX/NY/NZ535455
NX/NY/NZ134138134
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS927959305
D min/max/mean-3.0003.000-0.000

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Supplemental data

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Sample components

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Entire : 3D reconstruction and annotation of distinct secretory organelles...

EntireName: 3D reconstruction and annotation of distinct secretory organelles and cytoskeleton elements in the apical region of Toxoplasma gondii
Components
  • Cell: 3D reconstruction and annotation of distinct secretory organelles and cytoskeleton elements in the apical region of Toxoplasma gondii

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Supramolecule #1: 3D reconstruction and annotation of distinct secretory organelles...

SupramoleculeName: 3D reconstruction and annotation of distinct secretory organelles and cytoskeleton elements in the apical region of Toxoplasma gondii
type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Toxoplasma gondii (eukaryote)

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statecell

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Sample preparation

BufferpH: 7.2
Details: Hanks balanced salts solution, supplemented (HBSS): 1 mM MgCl2, 1 mM CaCl2, 10 mM NaHCO3, 20 mM HEPES, pH 7.2
GridModel: Homemade / Support film - Material: CARBON / Support film - topology: LACEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 25 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 295 K / Instrument: LEICA EM GP
Details3x10^6 cells/ml
SectioningOther: NO SECTIONING
Fiducial markerManufacturer: EMS / Diameter: 10 nm

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Specialist opticsPhase plate: VOLTA PHASE PLATE / Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 2.2 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 39000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Final reconstructionSoftware - Name: IMOD / Number images used: 41

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