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- EMDB-27565: Cryo-EM structure of nonmuscle gamma-actin -

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Basic information

Entry
Database: EMDB / ID: EMD-27565
TitleCryo-EM structure of nonmuscle gamma-actin
Map dataCryo-EM structure of nonmuscle gamma-actin
Sample
  • Organelle or cellular component: actin
    • Protein or peptide: Actin, cytoplasmic 2, N-terminally processed
  • Ligand: MAGNESIUM ION
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
Function / homology
Function and homology information


basal body patch / tight junction assembly / regulation of transepithelial transport / structural constituent of postsynaptic actin cytoskeleton / protein localization to bicellular tight junction / morphogenesis of a polarized epithelium / profilin binding / Formation of annular gap junctions / Gap junction degradation / dense body ...basal body patch / tight junction assembly / regulation of transepithelial transport / structural constituent of postsynaptic actin cytoskeleton / protein localization to bicellular tight junction / morphogenesis of a polarized epithelium / profilin binding / Formation of annular gap junctions / Gap junction degradation / dense body / Cell-extracellular matrix interactions / regulation of stress fiber assembly / Adherens junctions interactions / Sensory processing of sound by outer hair cells of the cochlea / Interaction between L1 and Ankyrins / Sensory processing of sound by inner hair cells of the cochlea / sarcomere organization / NuA4 histone acetyltransferase complex / regulation of synaptic vesicle endocytosis / apical junction complex / regulation of focal adhesion assembly / maintenance of blood-brain barrier / positive regulation of wound healing / myofibril / Recycling pathway of L1 / filamentous actin / calyx of Held / EPH-ephrin mediated repulsion of cells / RHO GTPases Activate WASPs and WAVEs / RHO GTPases activate IQGAPs / RHOBTB2 GTPase cycle / phagocytic vesicle / EPHB-mediated forward signaling / axonogenesis / actin filament / cell motility / RHO GTPases Activate Formins / Translocation of SLC2A4 (GLUT4) to the plasma membrane / FCGR3A-mediated phagocytosis / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / Signaling by high-kinase activity BRAF mutants / Schaffer collateral - CA1 synapse / MAP2K and MAPK activation / structural constituent of cytoskeleton / Regulation of actin dynamics for phagocytic cup formation / platelet aggregation / VEGFA-VEGFR2 Pathway / cellular response to type II interferon / Signaling by RAF1 mutants / Signaling by moderate kinase activity BRAF mutants / Paradoxical activation of RAF signaling by kinase inactive BRAF / Signaling downstream of RAS mutants / cell-cell junction / Signaling by BRAF and RAF1 fusions / Clathrin-mediated endocytosis / angiogenesis / blood microparticle / cytoskeleton / hydrolase activity / positive regulation of cell migration / axon / focal adhesion / synapse / ubiquitin protein ligase binding / positive regulation of gene expression / protein kinase binding / extracellular space / extracellular exosome / ATP binding / membrane / identical protein binding / nucleus / plasma membrane / cytosol / 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, cytoplasmic 2
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.38 Å
AuthorsArora AS / Huang HL / Heissler SM / Chinthalapudi K
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM143539 United States
CitationJournal: Elife / Year: 2023
Title: Structural insights into actin isoforms.
Authors: Amandeep S Arora / Hsiang-Ling Huang / Ramanpreet Singh / Yoshie Narui / Andrejus Suchenko / Tomoyuki Hatano / Sarah M Heissler / Mohan K Balasubramanian / Krishna Chinthalapudi /
Abstract: Actin isoforms organize into distinct networks that are essential for the normal function of eukaryotic cells. Despite a high level of sequence and structure conservation, subtle differences in their ...Actin isoforms organize into distinct networks that are essential for the normal function of eukaryotic cells. Despite a high level of sequence and structure conservation, subtle differences in their design principles determine the interaction with myosin motors and actin-binding proteins. Therefore, identifying how the structure of actin isoforms relates to function is important for our understanding of normal cytoskeletal physiology. Here, we report the high-resolution structures of filamentous skeletal muscle α-actin (3.37 Å), cardiac muscle α-actin (3.07 Å), ß-actin (2.99 Å), and γ-actin (3.38 Å) in the Mg·ADP state with their native post-translational modifications. The structures revealed isoform-specific conformations of the N-terminus that shift closer to the filament surface upon myosin binding, thereby establishing isoform-specific interfaces. Collectively, the structures of single-isotype, post-translationally modified bare skeletal muscle α-actin, cardiac muscle α-actin, ß-actin, and γ-actin reveal general principles, similarities, and differences between isoforms. They complement the repertoire of known actin structures and allow for a comprehensive understanding of in vitro and in vivo functions of actin isoforms.
History
DepositionJul 11, 2022-
Header (metadata) releaseApr 12, 2023-
Map releaseApr 12, 2023-
UpdateApr 12, 2023-
Current statusApr 12, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27565.png

Downloads & links

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Map

FileDownload / File: emd_27565.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM structure of nonmuscle gamma-actin
Voxel sizeX=Y=Z: 0.891 Å
Density
Contour LevelBy AUTHOR: 0.37
Minimum - Maximum-1.5182184 - 2.3246114
Average (Standard dev.)0.0051430548 (±0.095196694)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 228.096 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Cryo-EM structure of nonmuscle gamma-actin

Fileemd_27565_half_map_1.map
AnnotationCryo-EM structure of nonmuscle gamma-actin
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Cryo-EM structure of nonmuscle gamma-actin

Fileemd_27565_half_map_2.map
AnnotationCryo-EM structure of nonmuscle gamma-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 : actin

EntireName: actin
Components
  • Organelle or cellular component: actin
    • Protein or peptide: Actin, cytoplasmic 2, N-terminally processed
  • Ligand: MAGNESIUM ION
  • Ligand: ADENOSINE-5'-DIPHOSPHATE

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Supramolecule #1: actin

SupramoleculeName: actin / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Actin, cytoplasmic 2, N-terminally processed

MacromoleculeName: Actin, cytoplasmic 2, N-terminally processed / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 41.746625 KDa
Recombinant expressionOrganism: Pichia (fungus)
SequenceString: (ACE)EEEIAALVI DNGSGMCKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IE (HIC)GIVTNW DDMEKIWHHT FYNELRVAPE EHPVLLTEAP LNPKANREKM TQIMFETFNT PAMYVAIQAV LSLYASGRT TGIVMDSGDG ...String:
(ACE)EEEIAALVI DNGSGMCKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IE (HIC)GIVTNW DDMEKIWHHT FYNELRVAPE EHPVLLTEAP LNPKANREKM TQIMFETFNT PAMYVAIQAV LSLYASGRT TGIVMDSGDG VTHTVPIYEG YALPHAILRL DLAGRDLTDY LMKILTERGY SFTTTAEREI VRDIKEKLCY VALDFEQEMA TAASSSSLE KSYELPDGQV ITIGNERFRC PEALFQPSFL GMESCGIHET TFNSIMKCDV DIRKDLYANT VLSGGTTMYP G IADRMQKE ITALAPSTMK IKIIAPPERK YSVWIGGSIL ASLSTFQQMW ISKQEYDESG PSIVHRKCF

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Macromolecule #2: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 2 / Number of copies: 4 / 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: 4 / 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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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7
GridModel: C-flat / Material: GOLD / Support film - Material: CARBON / Support film - topology: CONTINUOUS
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 298 K / Instrument: LEICA EM GP

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.3 µm / Nominal magnification: 81000
Specialist opticsSpherical aberration corrector: Cs-corrected microscope / Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 2952 / Average electron dose: 65.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionDetails: Single Particle
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6djo

Initial angle assignmentType: NOT APPLICABLE / Software - Name: cryoSPARC
Final angle assignmentType: NOT APPLICABLE / Software - Name: cryoSPARC
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.38 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 1009372
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6djo


Chain - Chain ID: A
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 201
Output model

PDB-8dnf:
Cryo-EM structure of nonmuscle gamma-actin

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