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- EMDB-7007: CryoEM structure of Mical Oxidized Actin (Class 1) -

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Basic information

Entry
Database: EMDB / ID: EMD-7007
TitleCryoEM structure of Mical Oxidized Actin (Class 1)
Map data
Sample
  • Organelle or cellular component: Filamentous Actin Oxidized by Mical at M44 and M47
    • Protein or peptide: Actin, alpha skeletal muscle
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
Function / homology
Function and homology information


cytoskeletal motor activator activity / tropomyosin binding / myosin heavy chain binding / mesenchyme migration / troponin I binding / actin filament bundle / filamentous actin / actin filament bundle assembly / skeletal muscle thin filament assembly / striated muscle thin filament ...cytoskeletal motor activator activity / tropomyosin binding / myosin heavy chain binding / mesenchyme migration / troponin I binding / actin filament bundle / filamentous actin / actin filament bundle assembly / skeletal muscle thin filament assembly / striated muscle thin filament / skeletal muscle myofibril / 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) / Rabbit (rabbit)
Methodhelical reconstruction / cryo EM / Resolution: 3.9 Å
AuthorsGrintsevich EE / Ge P / Sawaya MR / Yesilyurt HG / Terman JR / Zhou ZH / Reisler E
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM071940 United States
National Science Foundation (NSF, United States)DMR-1548924 United States
CitationJournal: Nat Commun / Year: 2017
Title: Catastrophic disassembly of actin filaments via Mical-mediated oxidation.
Authors: Elena E Grintsevich / Peng Ge / Michael R Sawaya / Hunkar Gizem Yesilyurt / Jonathan R Terman / Z Hong Zhou / Emil Reisler /
Abstract: Actin filament assembly and disassembly are vital for cell functions. MICAL Redox enzymes are important post-translational effectors of actin that stereo-specifically oxidize actin's M44 and M47 ...Actin filament assembly and disassembly are vital for cell functions. MICAL Redox enzymes are important post-translational effectors of actin that stereo-specifically oxidize actin's M44 and M47 residues to induce cellular F-actin disassembly. Here we show that Mical-oxidized (Mox) actin can undergo extremely fast (84 subunits/s) disassembly, which depends on F-actin's nucleotide-bound state. Using near-atomic resolution cryoEM reconstruction and single filament TIRF microscopy we identify two dynamic and structural states of Mox-actin. Modeling actin's D-loop region based on our 3.9 Å cryoEM reconstruction suggests that oxidation by Mical reorients the side chain of M44 and induces a new intermolecular interaction of actin residue M47 (M47-O-T351). Site-directed mutagenesis reveals that this interaction promotes Mox-actin instability. Moreover, we find that Mical oxidation of actin allows for cofilin-mediated severing even in the presence of inorganic phosphate. Thus, in conjunction with cofilin, Mical oxidation of actin promotes F-actin disassembly independent of the nucleotide-bound state.
History
DepositionSep 1, 2017-
Header (metadata) releaseJan 17, 2018-
Map releaseJan 17, 2018-
UpdateNov 27, 2019-
Current statusNov 27, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.018
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.018
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6av9
  • Surface level: 0.018
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6av9
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_7007.png

Downloads & links

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Map

FileDownload / File: emd_7007.map.gz / Format: CCP4 / Size: 15.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.041 Å
Density
Contour LevelBy AUTHOR: 0.018 / Movie #1: 0.018
Minimum - Maximum-0.038082995 - 0.07752389
Average (Standard dev.)0.00001526440 (±0.006986138)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-80-80-80
Dimensions160160160
Spacing160160160
CellA=B=C: 166.56 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0411.0411.041
M x/y/z160160160
origin x/y/z0.0000.0000.000
length x/y/z166.560166.560166.560
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ240240240
MAP C/R/S123
start NC/NR/NS-80-80-80
NC/NR/NS160160160
D min/max/mean-0.0380.0780.000

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Supplemental data

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Sample components

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Entire : Filamentous Actin Oxidized by Mical at M44 and M47

EntireName: Filamentous Actin Oxidized by Mical at M44 and M47
Components
  • Organelle or cellular component: Filamentous Actin Oxidized by Mical at M44 and M47
    • Protein or peptide: Actin, alpha skeletal muscle
  • Ligand: ADENOSINE-5'-DIPHOSPHATE

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Supramolecule #1: Filamentous Actin Oxidized by Mical at M44 and M47

SupramoleculeName: Filamentous Actin Oxidized by Mical at M44 and M47 / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Oryctolagus cuniculus (rabbit)

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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: Rabbit (rabbit)
Molecular weightTheoretical: 42.141973 KDa
SequenceString: MCDEDETTAL VCDNGSGLVK AGFAGDDAPR AVFPSIVGRP RHQGV(SME)VG(SME)G QKDSYVGDEA QSKRGILTLK YP IE(HIC)GIIT NWDDMEKIWH HTFYNELRVA PEEHPTLLTE APLNPKANRE KMTQIMFETF NVPAMYVAIQ AVLSLYASG RTTGIVLDSG ...String:
MCDEDETTAL VCDNGSGLVK AGFAGDDAPR AVFPSIVGRP RHQGV(SME)VG(SME)G QKDSYVGDEA QSKRGILTLK YP IE(HIC)GIIT NWDDMEKIWH HTFYNELRVA PEEHPTLLTE APLNPKANRE KMTQIMFETF NVPAMYVAIQ AVLSLYASG RTTGIVLDSG DGVTHNVPIY EGYALPHAIM RLDLAGRDLT DYLMKILTER GYSFVTTAER EIVRDIKEKL CYVALDFENE MATAASSSS LEKSYELPDG QVITIGNERF RCPETLFQPS FIGMESAGIH ETTYNSIMKC DIDIRKDLYA NNVMSGGTTM Y PGIADRMQ 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 / Adenosine diphosphate

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statehelical array

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Sample preparation

BufferpH: 7.4
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV / Details: Blot Force 1, Blot time 4s.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal magnification: 130000
Specialist opticsEnergy filter - Name: Gatan Bio Quantum
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 4-20 / Average electron dose: 30.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware: (Name: CTFFIND (ver. 4), RELION (ver. 1.4))
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 1.4)
Final reconstructionApplied symmetry - Helical parameters - Δz: 28.042 Å
Applied symmetry - Helical parameters - Δ&Phi: -166.64 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.4) / Software - details: +IHRSR / Number images used: 102700
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: AB INITIO MODEL
Output model

PDB-6av9:
CryoEM structure of Mical Oxidized Actin (Class 1)

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