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- EMDB-3837: Cryo-EM structure of jasplakinolide-stabilized F-actin in complex... -

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Basic information

Entry
Database: EMDB / ID: EMD-3837
TitleCryo-EM structure of jasplakinolide-stabilized F-actin in complex with ADP-Pi
Map data
Sample
  • Complex: Filamentous jasplakinolide-stabilized alpha actin in complex with ADP-Pi
    • Protein or peptide: Actin, alpha skeletal muscle
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: PHOSPHATE ION
  • Ligand: (4~{R},7~{R},10~{S},13~{S},15~{E},19~{S})-10-(4-azanylbutyl)-4-(4-hydroxyphenyl)-7-(1~{H}-indol-3-ylmethyl)-8,13,15,19-tetramethyl-1-oxa-5,8,11-triazacyclononadec-15-ene-2,6,9,12-tetrone
  • Ligand: MAGNESIUM ION
KeywordsCytoskeleton / nucleotide states / filament stability / cell migration / STRUCTURAL PROTEIN
Function / homology
Function and homology information


cytoskeletal motor activator activity / myosin heavy chain binding / tropomyosin binding / actin filament bundle / troponin I binding / filamentous actin / mesenchyme migration / actin filament bundle assembly / skeletal muscle myofibril / striated muscle thin filament ...cytoskeletal motor activator activity / myosin heavy chain binding / tropomyosin binding / actin filament bundle / troponin I binding / filamentous actin / mesenchyme migration / actin filament bundle assembly / skeletal muscle myofibril / striated muscle thin filament / skeletal muscle thin filament assembly / actin monomer binding / skeletal muscle fiber development / stress fiber / titin binding / actin filament polymerization / filopodium / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / calcium-dependent protein binding / lamellipodium / cell body / hydrolase activity / protein domain specific binding / calcium ion binding / positive regulation of gene expression / magnesium ion binding / ATP binding / identical protein 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
Actin, alpha skeletal muscle
Similarity search - Component
Biological speciesOryctolagus cuniculus (rabbit)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsMerino F / Pospich S
Funding support Germany, 4 items
OrganizationGrant numberCountry
Max Planck Society Germany
State of Thuringia43- 5572-321-12040-12 Germany
European Union615984 Germany
Studienstiftung des deutschen Volkes Germany
CitationJournal: Nat Struct Mol Biol / Year: 2018
Title: Structural transitions of F-actin upon ATP hydrolysis at near-atomic resolution revealed by cryo-EM.
Authors: Felipe Merino / Sabrina Pospich / Johanna Funk / Thorsten Wagner / Florian Küllmer / Hans-Dieter Arndt / Peter Bieling / Stefan Raunser /
Abstract: The function of actin is coupled to the nucleotide bound to its active site. ATP hydrolysis is activated during polymerization; a delay between hydrolysis and inorganic phosphate (P) release results ...The function of actin is coupled to the nucleotide bound to its active site. ATP hydrolysis is activated during polymerization; a delay between hydrolysis and inorganic phosphate (P) release results in a gradient of ATP, ADP-P and ADP along actin filaments (F-actin). Actin-binding proteins can recognize F-actin's nucleotide state, using it as a local 'age' tag. The underlying mechanism is complex and poorly understood. Here we report six high-resolution cryo-EM structures of F-actin from rabbit skeletal muscle in different nucleotide states. The structures reveal that actin polymerization repositions the proposed catalytic base, His161, closer to the γ-phosphate. Nucleotide hydrolysis and P release modulate the conformational ensemble at the periphery of the filament, thus resulting in open and closed states, which can be sensed by coronin-1B. The drug-like toxin jasplakinolide locks F-actin in an open state. Our results demonstrate in detail how ATP hydrolysis links to F-actin's conformational dynamics and protein interaction.
History
DepositionAug 7, 2017-
Header (metadata) releaseDec 20, 2017-
Map releaseJun 13, 2018-
UpdateApr 9, 2025-
Current statusApr 9, 2025Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.032
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.032
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5ood
  • Surface level: 0.032
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-5ood
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3837.png

Downloads & links

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Map

FileDownload / File: emd_3837.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 288 pix.
= 313.92 Å		1.09 Å/pix.
x 288 pix.
= 313.92 Å		1.09 Å/pix.
x 288 pix.
= 313.92 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.09 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.032 / Movie #1: 0.032
Minimum - Maximum-0.07868639 - 0.1569087
Average (Standard dev.)0.00007516541 (±0.0062935404)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 313.92 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.091.091.09
M x/y/z288288288
origin x/y/z0.0000.0000.000
length x/y/z313.920313.920313.920
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS288288288
D min/max/mean-0.0790.1570.000

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Supplemental data

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Mask #1

Fileemd_3837_msk_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_3837_half_map_1.map
Projections & Slices
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Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: #2

Fileemd_3837_half_map_2.map
Projections & Slices
AxesZYX

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Sample components

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Entire : Filamentous jasplakinolide-stabilized alpha actin in complex with...

EntireName: Filamentous jasplakinolide-stabilized alpha actin in complex with ADP-Pi
Components
  • Complex: Filamentous jasplakinolide-stabilized alpha actin in complex with ADP-Pi
    • Protein or peptide: Actin, alpha skeletal muscle
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: PHOSPHATE ION
  • Ligand: (4~{R},7~{R},10~{S},13~{S},15~{E},19~{S})-10-(4-azanylbutyl)-4-(4-hydroxyphenyl)-7-(1~{H}-indol-3-ylmethyl)-8,13,15,19-tetramethyl-1-oxa-5,8,11-triazacyclononadec-15-ene-2,6,9,12-tetrone
  • Ligand: MAGNESIUM ION

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Supramolecule #1: Filamentous jasplakinolide-stabilized alpha actin in complex with...

SupramoleculeName: Filamentous jasplakinolide-stabilized alpha actin in complex with ADP-Pi
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Oryctolagus cuniculus (rabbit) / Tissue: skeletal muscle

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Macromolecule #1: Actin, alpha skeletal muscle

MacromoleculeName: Actin, alpha skeletal muscle / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Oryctolagus cuniculus (rabbit) / Tissue: skeletal muscle
Molecular weightTheoretical: 41.875633 KDa
SequenceString: DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IE(HIC)GII TNW DDMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSG DG VTHNVPIYEG ...String:
DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IE(HIC)GII TNW DDMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSG DG VTHNVPIYEG YALPHAIMRL DLAGRDLTDY LMKILTERGY SFVTTAEREI VRDIKEKLCY VALDFENEMA TAASSSSL E KSYELPDGQV ITIGNERFRC PETLFQPSFI GMESAGIHET TYNSIMKCDI DIRKDLYANN VMSGGTTMYP GIADRMQKE ITALAPSTMK IKIIAPPERK YSVWIGGSIL ASLSTFQQMW ITKQEYDEAG PSIVHRKCF

UniProtKB: Actin, alpha skeletal muscle

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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

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Macromolecule #3: PHOSPHATE ION

MacromoleculeName: PHOSPHATE ION / type: ligand / ID: 3 / Number of copies: 5 / Formula: PO4
Molecular weightTheoretical: 94.971 Da
Chemical component information

ChemComp-PO4:
PHOSPHATE ION

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Macromolecule #4: (4~{R},7~{R},10~{S},13~{S},15~{E},19~{S})-10-(4-azanylbutyl)-4-(4...

MacromoleculeName: (4~{R},7~{R},10~{S},13~{S},15~{E},19~{S})-10-(4-azanylbutyl)-4-(4-hydroxyphenyl)-7-(1~{H}-indol-3-ylmethyl)-8,13,15,19-tetramethyl-1-oxa-5,8,11-triazacyclononadec-15-ene-2,6,9,12-tetrone
type: ligand / ID: 4 / Number of copies: 5 / Formula: 9ZK
Molecular weightTheoretical: 673.841 Da
Chemical component information

ChemComp-9ZK:
(4~{R},7~{R},10~{S},13~{S},15~{E},19~{S})-10-(4-azanylbutyl)-4-(4-hydroxyphenyl)-7-(1~{H}-indol-3-ylmethyl)-8,13,15,19-tetramethyl-1-oxa-5,8,11-triazacyclononadec-15-ene-2,6,9,12-tetrone

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Macromolecule #5: MAGNESIUM ION

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

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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:
ConcentrationFormulaName
5.0 mMC4H11NO3TRIS
2.0 mMNaN3Sodium azide
1.0 mMC4H10O2S2DTT
100.0 mMKClPotassium chloride
2.0 mMMgCl2Magnesium chloride
9.0 % (v/v)C2H6OSDMSO

Details: 5 mM Tris pH 7.5, 2 mM NaN3, 1 mM DTT, 100 mM KCl and 2 mM MgCl2, 9.0 %(v/v) DMSO
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 298 K / Instrument: GATAN CRYOPLUNGE 3
Details: Manual backside blotting using Whatman filter paper No.5..
DetailsRise 27.39 A, Twist -166.41 degrees

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsSpherical aberration corrector: Cs-corrected microscope / Energy filter - Name: GIF Quantum ER
DetailsCs-corrected microscope
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-15 / Number real images: 724 / Average exposure time: 15.0 sec. / Average electron dose: 78.0 e/Å2 / Details: Images were binned twice during motion correction.
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 154223
Startup modelType of model: INSILICO MODEL
In silico model: Initial model was generated from PDB 5JLF and filtered to 20 A using SPARX.
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.4) / Number images used: 123716
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 1.4)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 1.4)

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Atomic model buiding 1

DetailsRosetta iterative refinement was combined with MDFF.
RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-5ood:
Cryo-EM structure of jasplakinolide-stabilized F-actin in complex with ADP-Pi

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