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- EMDB-41584: Actin 1 from T. gondii in filaments bound to MgADP and jasplakinolide -

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Basic information

Entry
Database: EMDB / ID: EMD-41584
TitleActin 1 from T. gondii in filaments bound to MgADP and jasplakinolide
Map dataActin 1 from T. gondii in filaments with MgADP and jasplakinolide; Density modified map from helical reconstruction
Sample
  • Complex: Filament of Actin 1 from T. gondii with MgADP and jasplakinolide
    • Protein or peptide: Actin
  • Ligand: MAGNESIUM ION
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: Jasplakinolide
KeywordsActin / cytoskeleton / toxoplasmosis / STRUCTURAL PROTEIN
Function / homology
Function and homology information


Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / cytoskeleton / hydrolase activity / ATP binding / cytoplasm
Similarity search - Function
Actins signature 1. / Actin, conserved site / Actins signature 2. / Actin/actin-like conserved site / Actins and actin-related proteins signature. / Actin / Actin family / Actin / ATPase, nucleotide binding domain
Similarity search - Domain/homology
Biological speciesToxoplasma gondii (eukaryote)
Methodhelical reconstruction / cryo EM / Resolution: 3.0 Å
AuthorsHvorecny KL / Sladewski TE / Heaslip AT / Kollman JM
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)F32AI145111 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM138316 United States
CitationJournal: Nat Commun / Year: 2024
Title: Toxoplasma gondii actin filaments are tuned for rapid disassembly and turnover.
Authors: Kelli L Hvorecny / Thomas E Sladewski / Enrique M De La Cruz / Justin M Kollman / Aoife T Heaslip /
Abstract: The cytoskeletal protein actin plays a critical role in the pathogenicity of the intracellular parasite, Toxoplasma gondii, mediating invasion and egress, cargo transport, and organelle inheritance. ...The cytoskeletal protein actin plays a critical role in the pathogenicity of the intracellular parasite, Toxoplasma gondii, mediating invasion and egress, cargo transport, and organelle inheritance. Advances in live cell imaging have revealed extensive filamentous actin networks in the Apicomplexan parasite, but there are conflicting data regarding the biochemical and biophysical properties of Toxoplasma actin. Here, we imaged the in vitro assembly of individual Toxoplasma actin filaments in real time, showing that native, unstabilized filaments grow tens of microns in length. Unlike skeletal muscle actin, Toxoplasma filaments intrinsically undergo rapid treadmilling due to a high critical concentration, fast monomer dissociation, and rapid nucleotide exchange. Cryo-EM structures of jasplakinolide-stabilized and native (i.e. unstabilized) filaments show an architecture like skeletal actin, with differences in assembly contacts in the D-loop that explain the dynamic nature of the filament, likely a conserved feature of Apicomplexan actin. This work demonstrates that evolutionary changes at assembly interfaces can tune the dynamic properties of actin filaments without disrupting their conserved structure.
History
DepositionAug 9, 2023-
Header (metadata) releaseMar 6, 2024-
Map releaseMar 6, 2024-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41584.png

Downloads & links

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Map

FileDownload / File: emd_41584.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationActin 1 from T. gondii in filaments with MgADP and jasplakinolide; Density modified map from helical reconstruction
Voxel sizeX=Y=Z: 0.89 Å
Density
Contour LevelBy AUTHOR: 1.75
Minimum - Maximum-8.917244 - 16.445307
Average (Standard dev.)-0.000009252803 (±0.34443358)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 341.76 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Three protomers of Actin 1 from T. gondii...

Fileemd_41584_additional_1.map
AnnotationThree protomers of Actin 1 from T. gondii in filaments with MgADP and jasplakinolide; Half map from local refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Three protomers of Actin 1 from T. gondii...

Fileemd_41584_additional_2.map
AnnotationThree protomers of Actin 1 from T. gondii in filaments with MgADP and jasplakinolide; Density modified map from local refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Three protomers of Actin 1 from T. gondii...

Fileemd_41584_additional_3.map
AnnotationThree protomers of Actin 1 from T. gondii in filaments with MgADP and jasplakinolide; Half map from local refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Actin 1 from T. gondii in filaments with...

Fileemd_41584_half_map_1.map
AnnotationActin 1 from T. gondii in filaments with MgADP and jasplakinolide; Half map from helical reconstruction
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Actin 1 from T. gondii in filaments with...

Fileemd_41584_half_map_2.map
AnnotationActin 1 from T. gondii in filaments with MgADP and jasplakinolide; Half map from helical reconstruction
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Filament of Actin 1 from T. gondii with MgADP and jasplakinolide

EntireName: Filament of Actin 1 from T. gondii with MgADP and jasplakinolide
Components
  • Complex: Filament of Actin 1 from T. gondii with MgADP and jasplakinolide
    • Protein or peptide: Actin
  • Ligand: MAGNESIUM ION
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: Jasplakinolide

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Supramolecule #1: Filament of Actin 1 from T. gondii with MgADP and jasplakinolide

SupramoleculeName: Filament of Actin 1 from T. gondii with MgADP and jasplakinolide
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Actin protomers from T. gondii assemble into a two-start helix, stabilized by jasplakinolide
Source (natural)Organism: Toxoplasma gondii (eukaryote) / Strain: Sf9

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Macromolecule #1: Actin

MacromoleculeName: Actin / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
EC number: Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement
Source (natural)Organism: Toxoplasma gondii (eukaryote)
Molecular weightTheoretical: 41.956711 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MADEEVQALV VDNGSGNVKA GVAGDDAPRA VFPSIVGKPK NPGIMVGMEE KDCYVGDEAQ SKRGILTLKY PIEHGIVTNW DDMEKIWHH TFYNELRVAP EEHPVLLTEA PLNPKANRER MTQIMFETFN VPAMYVAIQA VLSLYSSGRT TGIVLDSGDG V SHTVPIYE ...String:
MADEEVQALV VDNGSGNVKA GVAGDDAPRA VFPSIVGKPK NPGIMVGMEE KDCYVGDEAQ SKRGILTLKY PIEHGIVTNW DDMEKIWHH TFYNELRVAP EEHPVLLTEA PLNPKANRER MTQIMFETFN VPAMYVAIQA VLSLYSSGRT TGIVLDSGDG V SHTVPIYE GYALPHAIMR LDLAGRDLTE YMMKILHERG YGFTTSAEKE IVRDIKEKLC YIALDFDEEM KAAEDSSDIE KS YELPDGN IITVGNERFR CPEALFQPSF LGKEAAGVHR TTFDSIMKCD VDIRKDLYGN VVLSGGTTMY EGIGERLTKE LTS LAPSTM KIKVVAPPER KYSVWIGGSI LSSLSTFQQM WITKEEYDES GPSIVHRKCF

UniProtKB: Actin

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Macromolecule #2: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 2 / Number of copies: 3 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #3: ADENOSINE-5'-DIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 3 / 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 #4: Jasplakinolide

MacromoleculeName: Jasplakinolide / type: ligand / ID: 4 / Number of copies: 3 / Formula: 9UE
Molecular weightTheoretical: 709.67 Da
Chemical component information

ChemComp-9UE:
Jasplakinolide

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 8
GridModel: C-flat-1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER

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Electron microscopy

MicroscopeTFS GLACIOS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.75 µm / Nominal magnification: 45000
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Detector mode: SUPER-RESOLUTION / Average exposure time: 5.0 sec. / Average electron dose: 50.0 e/Å2

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Image processing

Startup modelType of model: NONE
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2)
Final reconstructionApplied symmetry - Helical parameters - Δz: 28.1 Å
Applied symmetry - Helical parameters - Δ&Phi: -167 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: PHENIX (ver. 1.20) / Number images used: 2682453

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Atomic model buiding 1

RefinementProtocol: FLEXIBLE FIT
Output model

PDB-8trn:
Actin 1 from T. gondii in filaments bound to MgADP and jasplakinolide

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