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- EMDB-3805: Cryo-EM structure of jasplakinolide-stabilized malaria parasite F... -

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Basic information

Entry
Database: EMDB / ID: EMD-3805
TitleCryo-EM structure of jasplakinolide-stabilized malaria parasite F-actin at near-atomic resolution
Map data
Sample
  • Complex: Plasmodium falciparum actin 1 filament stabilized by jasplakinolide
    • Protein or peptide: Actin-1
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: Jasplakinolide
Function / homology
Function and homology information


actin polymerization-dependent cell-to-cell migration in host / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / structural constituent of cytoskeleton / hydrolase activity / ATP binding / nucleus / 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 speciesPlasmodium falciparum HB3 (eukaryote) / Plasmodium falciparum (isolate HB3) (eukaryote)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsPospich S / Kumpula E-P / von der Ecken J / Vahokoski J / Kursula I / Raunser S
Funding support Germany, Finland, 4 items
OrganizationGrant numberCountry
Max Planck Society Germany
European Union615984
Academy of Finland257537, 265112, 292718 Finland
Emil Aaltonen, Sigrid Juselius, and Jane and Aatos Erkko foundations
CitationJournal: Proc Natl Acad Sci U S A / Year: 2017
Title: Near-atomic structure of jasplakinolide-stabilized malaria parasite F-actin reveals the structural basis of filament instability.
Authors: Sabrina Pospich / Esa-Pekka Kumpula / Julian von der Ecken / Juha Vahokoski / Inari Kursula / Stefan Raunser /
Abstract: During their life cycle, apicomplexan parasites, such as the malaria parasite , use actomyosin-driven gliding motility to move and invade host cells. For this process, actin filament length and ...During their life cycle, apicomplexan parasites, such as the malaria parasite , use actomyosin-driven gliding motility to move and invade host cells. For this process, actin filament length and stability are temporally and spatially controlled. In contrast to canonical actin, actin 1 (Act1) does not readily polymerize into long, stable filaments. The structural basis of filament instability, which plays a pivotal role in host cell invasion, and thus infectivity, is poorly understood, largely because high-resolution structures of Act1 filaments were missing. Here, we report the near-atomic structure of jasplakinolide (JAS)-stabilized Act1 filaments determined by electron cryomicroscopy. The general filament architecture is similar to that of mammalian F-actin. The high resolution of the structure allowed us to identify small but important differences at inter- and intrastrand contact sites, explaining the inherent instability of apicomplexan actin filaments. JAS binds at regular intervals inside the filament to three adjacent actin subunits, reinforcing filament stability by hydrophobic interactions. Our study reveals the high-resolution structure of a small molecule bound to F-actin, highlighting the potential of electron cryomicroscopy for structure-based drug design. Furthermore, our work serves as a strong foundation for understanding the structural design and evolution of actin filaments and their function in motility and host cell invasion of apicomplexan parasites.
History
DepositionJul 13, 2017-
Header (metadata) releaseJul 26, 2017-
Map releaseSep 27, 2017-
UpdateNov 9, 2022-
Current statusNov 9, 2022Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.09
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.09
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5ogw
  • Surface level: 0.09
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-5ogw
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3805.png

Downloads & links

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Map

FileDownload / File: emd_3805.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.14 Å
Density
Contour LevelBy AUTHOR: 0.09 / Movie #1: 0.09
Minimum - Maximum-0.15085225 - 0.3457834
Average (Standard dev.)0.0014830945 (±0.013865351)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 291.84 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.141.141.14
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z291.840291.840291.840
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.1510.3460.001

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Supplemental data

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Mask #1

Fileemd_3805_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_3805_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_3805_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Plasmodium falciparum actin 1 filament stabilized by jasplakinolide

EntireName: Plasmodium falciparum actin 1 filament stabilized by jasplakinolide
Components
  • Complex: Plasmodium falciparum actin 1 filament stabilized by jasplakinolide
    • Protein or peptide: Actin-1
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: Jasplakinolide

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Supramolecule #1: Plasmodium falciparum actin 1 filament stabilized by jasplakinolide

SupramoleculeName: Plasmodium falciparum actin 1 filament stabilized by jasplakinolide
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: Filament
Source (natural)Organism: Plasmodium falciparum HB3 (eukaryote)
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)

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Macromolecule #1: Actin-1

MacromoleculeName: Actin-1 / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Plasmodium falciparum (isolate HB3) (eukaryote) / Strain: isolate HB3
Molecular weightTheoretical: 42.047676 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: GAMGEEDVQA LVVDNGSGNV KAGVAGDDAP RSVFPSIVGR PKNPGIMVGM EEKDAFVGDE AQTKRGILTL KYPIEHGIVT NWDDMEKIW HHTFYNELRA APEEHPVLLT EAPLNPKGNR ERMTQIMFES FNVPAMYVAI QAVLSLYSSG RTTGIVLDSG D GVSHTVPI ...String:
GAMGEEDVQA LVVDNGSGNV KAGVAGDDAP RSVFPSIVGR PKNPGIMVGM EEKDAFVGDE AQTKRGILTL KYPIEHGIVT NWDDMEKIW HHTFYNELRA APEEHPVLLT EAPLNPKGNR ERMTQIMFES FNVPAMYVAI QAVLSLYSSG RTTGIVLDSG D GVSHTVPI YEGYALPHAI MRLDLAGRDL TEYLMKILHE RGYGFSTSAE KEIVRDIKEK LCYIALNFDE EMKTSEQSSD IE KSYELPD GNIITVGNER FRCPEALFQP SFLGKEAAGI HTTTFNSIKK CDVDIRKDLY GNIVLSGGTT MYEGIGERLT RDI TTLAPS TMKIKVVAPP ERKYSVWIGG SILSSLSTFQ QMWITKEEYD ESGPSIVHRK CF

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Macromolecule #2: ADENOSINE-5'-DIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 2 / Number of copies: 5 / 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 #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 5 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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
Processingsingle particle reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.5
Component:
ConcentrationFormula
10.0 mMHEPES
0.2 mMCaCl2
50.0 mMKCl
5.0 mMDTT

Details: 10 mM HEPES pH 7.5, 0.2 mM CaCl2, 50 mM KCl, 4 mM MgCl2, 5 mM DTT and 0.5 mM ATP. JAS was added at a 1:1 molar ratio during polyermization.
GridModel: C-flat-2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 97 % / Chamber temperature: 298 K / Instrument: GATAN CRYOPLUNGE 3
Details: Sample (2 uL of JAS-stabilized F-actin solution) was applied to a glow-discharged holey carbon grid, incubated for 30 s and manually blotted for 4 s from the backside with filter paper..
DetailsTwist (degree) 167.5 Rise (A) 27.4

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.7 µm / Nominal defocus min: 0.8 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
DetailsCs corrected microscope
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Digitization - Frames/image: 1-4 / Number real images: 1634 / Average exposure time: 1.5 sec. / Average electron dose: 110.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 144058
CTF correctionSoftware: (Name: CTFFIND (ver. 4.0.7), RELION (ver. 1.4))
Startup modelType of model: INSILICO MODEL
Details: An inital model was generated by first fitting a homology model into a previously obtained map at 7 A resolution and afterwards creating an electron density map from the model and filtering it to 20 A.
Initial angle assignmentType: PROJECTION MATCHING
Projection matching processing - Angular sampling: 3.7 degrees
Software - Name: RELION (ver. 1.4)
Final angle assignmentType: PROJECTION MATCHING
Projection matching processing - Angular sampling: 0.9 degrees
Software - Name: RELION (ver. 1.4)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.4) / Number images used: 140716

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