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- EMDB-27114: Helical ADP-F-actin -

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Basic information

Entry
Database: EMDB / ID: EMD-27114
TitleHelical ADP-F-actin
Map dataFinal refine3d map, unsharpened, unboxed, unmasked
Sample
  • Complex: Helical F-actin, ADP nucleotide state
    • Protein or peptide: Actin, alpha skeletal muscle
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: water
Function / homology
Function and homology information


Striated Muscle Contraction / skeletal muscle thin filament assembly / striated muscle thin filament / stress fiber / skeletal muscle fiber development / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / hydrolase activity / ATP binding
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 speciesGallus gallus (chicken) / chicken (chicken)
Methodhelical reconstruction / cryo EM / Resolution: 2.43 Å
AuthorsReynolds MJ / Alushin GM
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM141044 United States
CitationJournal: Nature / Year: 2022
Title: Bending forces and nucleotide state jointly regulate F-actin structure.
Authors: Matthew J Reynolds / Carla Hachicho / Ayala G Carl / Rui Gong / Gregory M Alushin /
Abstract: ATP-hydrolysis-coupled actin polymerization is a fundamental mechanism of cellular force generation. In turn, force and actin filament (F-actin) nucleotide state regulate actin dynamics by tuning F- ...ATP-hydrolysis-coupled actin polymerization is a fundamental mechanism of cellular force generation. In turn, force and actin filament (F-actin) nucleotide state regulate actin dynamics by tuning F-actin's engagement of actin-binding proteins through mechanisms that are unclear. Here we show that the nucleotide state of actin modulates F-actin structural transitions evoked by bending forces. Cryo-electron microscopy structures of ADP-F-actin and ADP-P-F-actin with sufficient resolution to visualize bound solvent reveal intersubunit interfaces bridged by water molecules that could mediate filament lattice flexibility. Despite extensive ordered solvent differences in the nucleotide cleft, these structures feature nearly identical lattices and essentially indistinguishable protein backbone conformations that are unlikely to be discriminable by actin-binding proteins. We next introduce a machine-learning-enabled pipeline for reconstructing bent filaments, enabling us to visualize both continuous structural variability and side-chain-level detail. Bent F-actin structures reveal rearrangements at intersubunit interfaces characterized by substantial alterations of helical twist and deformations in individual protomers, transitions that are distinct in ADP-F-actin and ADP-P-F-actin. This suggests that phosphate rigidifies actin subunits to alter the bending structural landscape of F-actin. As bending forces evoke nucleotide-state dependent conformational transitions of sufficient magnitude to be detected by actin-binding proteins, we propose that actin nucleotide state can serve as a co-regulator of F-actin mechanical regulation.
History
DepositionMay 26, 2022-
Header (metadata) releaseSep 7, 2022-
Map releaseSep 7, 2022-
UpdateNov 23, 2022-
Current statusNov 23, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27114.png

Downloads & links

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Map

FileDownload / File: emd_27114.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFinal refine3d map, unsharpened, unboxed, unmasked
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.03 Å/pix.
x 512 pix.
= 527.36 Å		1.03 Å/pix.
x 512 pix.
= 527.36 Å		1.03 Å/pix.
x 512 pix.
= 527.36 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.03 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.025
Minimum - Maximum-0.014195239 - 0.085992336
Average (Standard dev.)3.364915e-06 (±0.0014277772)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 527.36 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_27114_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Mapboxed, 4x upsampled, density modified map to help model building

Fileemd_27114_additional_1.map
AnnotationMapboxed, 4x upsampled, density modified map to help model building
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 2, helical ADP F-actin

Fileemd_27114_half_map_1.map
AnnotationHalf map 2, helical ADP F-actin
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1, helical ADP F-actin

Fileemd_27114_half_map_2.map
AnnotationHalf map 1, helical ADP F-actin
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Helical F-actin, ADP nucleotide state

EntireName: Helical F-actin, ADP nucleotide state
Components
  • Complex: Helical F-actin, ADP nucleotide state
    • Protein or peptide: Actin, alpha skeletal muscle
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: water

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Supramolecule #1: Helical F-actin, ADP nucleotide state

SupramoleculeName: Helical F-actin, ADP nucleotide state / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Helically symmetric filamentous actin in the aged ADP state
Source (natural)Organism: Gallus gallus (chicken)
Molecular weightTheoretical: 15.1 kDa/nm

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Macromolecule #1: Actin, alpha skeletal muscle

MacromoleculeName: Actin, alpha skeletal muscle / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: chicken (chicken)
Molecular weightTheoretical: 42.109973 KDa
SequenceString: MCDEDETTAL VCDNGSGLVK AGFAGDDAPR AVFPSIVGRP RHQGVMVGMG QKDSYVGDEA QSKRGILTLK YPIE(HIC)G IIT NWDDMEKIWH HTFYNELRVA PEEHPTLLTE APLNPKANRE KMTQIMFETF NVPAMYVAIQ AVLSLYASGR TTGIVLD SG DGVTHNVPIY ...String:
MCDEDETTAL VCDNGSGLVK AGFAGDDAPR AVFPSIVGRP RHQGVMVGMG QKDSYVGDEA QSKRGILTLK YPIE(HIC)G IIT NWDDMEKIWH HTFYNELRVA PEEHPTLLTE APLNPKANRE KMTQIMFETF NVPAMYVAIQ AVLSLYASGR TTGIVLD SG DGVTHNVPIY EGYALPHAIM RLDLAGRDLT DYLMKILTER GYSFVTTAER EIVRDIKEKL CYVALDFENE MATAASSS S LEKSYELPDG QVITIGNERF RCPETLFQPS FIGMESAGIH ETTYNSIMKC DIDIRKDLYA NNVMSGGTTM YPGIADRMQ KEITALAPST MKIKIIAPPE RKYSVWIGGS ILASLSTFQQ MWITKQEYDE AGPSIVHRKC F

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Macromolecule #2: ADENOSINE-5'-DIPHOSPHATE

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

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

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 432 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-40 / Number grids imaged: 1 / Average exposure time: 10.0 sec. / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 4.0 µm / Nominal defocus min: 2.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionNumber classes used: 1
Applied symmetry - Helical parameters - Δz: 28.0687 Å
Applied symmetry - Helical parameters - Δ&Phi: -166.694 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 2.43 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 219569
CTF correctionSoftware - Name: CTFFIND (ver. 4.0)
Segment selectionNumber selected: 220462
Startup modelType of model: NONE
Details: Ab initio reconstruction as implemented in cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
FSC plot (resolution estimation)

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