[English] 日本語
Yorodumi
- EMDB-6451: Unsharpened cryo-EM reconstruction of the GFP-E892-Vt-actin inter... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-6451
TitleUnsharpened cryo-EM reconstruction of the GFP-E892-Vt-actin interface for difference map calculation
Map dataUnsharpened reconstruction of N-terminal GFP fusion to VT residues 892-1061 bound to actin for difference map calculation
Sample
  • Sample: GFP-E892-Vt bound to F-actin
  • Protein or peptide: skeletal muscle actin
  • Protein or peptide: Vinculin tail domain, residues 892-1061
Keywordsactin / vinculin / cell migration / adhesion / mechanosensation / cytoskeleton
Function / homology
Function and homology information


muscle tendon junction / Platelet degranulation / Smooth Muscle Contraction / regulation of protein localization to adherens junction / podosome ring / outer dense plaque of desmosome / inner dense plaque of desmosome / terminal web / epithelial cell-cell adhesion / zonula adherens ...muscle tendon junction / Platelet degranulation / Smooth Muscle Contraction / regulation of protein localization to adherens junction / podosome ring / outer dense plaque of desmosome / inner dense plaque of desmosome / terminal web / epithelial cell-cell adhesion / zonula adherens / dystroglycan binding / vinculin binding / muscle alpha-actinin binding / MAP2K and MAPK activation / alpha-catenin binding / fascia adherens / cell-cell contact zone / costamere / apical junction assembly / regulation of establishment of endothelial barrier / adherens junction assembly / axon extension / protein localization to cell surface / lamellipodium assembly / cytoskeletal motor activator activity / regulation of focal adhesion assembly / tropomyosin binding / myosin heavy chain binding / mesenchyme migration / troponin I binding / actin filament bundle / alpha-actinin binding / filamentous actin / actin filament bundle assembly / skeletal muscle thin filament assembly / brush border / striated muscle thin filament / skeletal muscle myofibril / actin monomer binding / skeletal muscle fiber development / stress fiber / regulation of cell migration / titin binding / actin filament polymerization / filopodium / cell projection / actin filament / Neutrophil degranulation / morphogenesis of an epithelium / adherens junction / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / sarcolemma / neuromuscular junction / Z disc / beta-catenin binding / calcium-dependent protein binding / actin filament binding / cell-cell junction / actin cytoskeleton / lamellipodium / cell body / scaffold protein binding / mitochondrial inner membrane / cytoskeleton / hydrolase activity / cell adhesion / cadherin binding / protein domain specific binding / focal adhesion / ubiquitin protein ligase binding / calcium ion binding / structural molecule activity / positive regulation of gene expression / magnesium ion binding / protein homodimerization activity / protein-containing complex / ATP binding / identical protein binding / plasma membrane / cytoplasm
Similarity search - Function
Vinculin repeated domain signature. / Vinculin / Vinculin, conserved site / Vinculin family talin-binding region signature. / Vinculin/alpha-catenin / Vinculin family / Alpha-catenin/vinculin-like superfamily / Actins signature 1. / Actin, conserved site / Actins signature 2. ...Vinculin repeated domain signature. / Vinculin / Vinculin, conserved site / Vinculin family talin-binding region signature. / Vinculin/alpha-catenin / Vinculin family / Alpha-catenin/vinculin-like superfamily / 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
Vinculin / Actin, alpha skeletal muscle
Similarity search - Component
Biological speciesOryctolagus cuniculus (rabbit) / Gallus gallus (chicken)
Methodhelical reconstruction / cryo EM / Resolution: 16.4 Å
AuthorsKim LY / Thompson PM / Lee HT / Pershad M / Campbell SL / Alushin GM
CitationJournal: J Mol Biol / Year: 2016
Title: The Structural Basis of Actin Organization by Vinculin and Metavinculin.
Authors: Laura Y Kim / Peter M Thompson / Hyunna T Lee / Mihir Pershad / Sharon L Campbell / Gregory M Alushin /
Abstract: Vinculin is an essential adhesion protein that links membrane-bound integrin and cadherin receptors through their intracellular binding partners to filamentous actin, facilitating mechanotransduction. ...Vinculin is an essential adhesion protein that links membrane-bound integrin and cadherin receptors through their intracellular binding partners to filamentous actin, facilitating mechanotransduction. Here we present an 8.5-Å-resolution cryo-electron microscopy reconstruction and pseudo-atomic model of the vinculin tail (Vt) domain bound to F-actin. Upon actin engagement, the N-terminal "strap" and helix 1 are displaced from the Vt helical bundle to mediate actin bundling. We find that an analogous conformational change also occurs in the H1' helix of the tail domain of metavinculin (MVt) upon actin binding, a muscle-specific splice isoform that suppresses actin bundling by Vt. These data support a model in which metavinculin tunes the actin bundling activity of vinculin in a tissue-specific manner, providing a mechanistic framework for understanding metavinculin mutations associated with hereditary cardiomyopathies.
History
DepositionSep 3, 2015-
Header (metadata) releaseSep 23, 2015-
Map releaseNov 4, 2015-
UpdateFeb 17, 2016-
Current statusFeb 17, 2016Processing site: RCSB / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3.5
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 3.5
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_6451.png

Downloads & links

-
Map

FileDownload / File: emd_6451.map.gz / Format: CCP4 / Size: 62.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationUnsharpened reconstruction of N-terminal GFP fusion to VT residues 892-1061 bound to actin for difference map calculation
Voxel sizeX=Y=Z: 2.18 Å
Density
Contour LevelBy AUTHOR: 3.5 / Movie #1: 3.5
Minimum - Maximum-3.69131851 - 13.673188209999999
Average (Standard dev.)0.0 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-128-128-128
Dimensions256256256
Spacing256256256
CellA=B=C: 558.08 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.182.182.18
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z558.080558.080558.080
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-128-128-128
NC/NR/NS256256256
D min/max/mean-3.69113.6730.000

-
Supplemental data

-
Sample components

-
Entire : GFP-E892-Vt bound to F-actin

EntireName: GFP-E892-Vt bound to F-actin
Components
  • Sample: GFP-E892-Vt bound to F-actin
  • Protein or peptide: skeletal muscle actin
  • Protein or peptide: Vinculin tail domain, residues 892-1061

-
Supramolecule #1000: GFP-E892-Vt bound to F-actin

SupramoleculeName: GFP-E892-Vt bound to F-actin / type: sample / ID: 1000
Details: Helical filament with one vinculin tail domain bound per actin protomer
Oligomeric state: One vinculin tail domain per actin protomer
Number unique components: 2

-
Macromolecule #1: skeletal muscle actin

MacromoleculeName: skeletal muscle actin / type: protein_or_peptide / ID: 1 / Name.synonym: actin / Oligomeric state: helical filament / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Oryctolagus cuniculus (rabbit) / synonym: Rabbit / Tissue: Muscle / Location in cell: Cytoplasm, cytoskeleton
Molecular weightTheoretical: 41.8 KDa
SequenceUniProtKB: Actin, alpha skeletal muscle

-
Macromolecule #2: Vinculin tail domain, residues 892-1061

MacromoleculeName: Vinculin tail domain, residues 892-1061 / type: protein_or_peptide / ID: 2 / Name.synonym: GFP-E892-Vt / Details: N-terminal GFP fusion / Oligomeric state: helical / Recombinant expression: Yes
Source (natural)Organism: Gallus gallus (chicken) / synonym: Chicken / Organelle: Focal adhesion / Location in cell: Cytoplasmic
Molecular weightTheoretical: 46 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria) / Recombinant strain: BL21(DE3) / Recombinant plasmid: H6-msfGFP (Addgene #29725)
SequenceUniProtKB: Vinculin

-
Experimental details

-
Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

-
Sample preparation

Concentration0.0125 mg/mL
BufferpH: 7 / Details: 50 mM KCl, 1 mM MgCl2, 1mM EGTA, 10 mM imidazole
GridDetails: 200 mesh 1.2 / 1.3 C-flat
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Instrument: LEICA EM GP
Method: 3 microliters of 0.3 micromolar actin was applied to the grid and incubated for 60 seconds at 25 degrees C. 3 microliters of 10 micromolar GFP-E892-Vt was then applied and incubated for 60 ...Method: 3 microliters of 0.3 micromolar actin was applied to the grid and incubated for 60 seconds at 25 degrees C. 3 microliters of 10 micromolar GFP-E892-Vt was then applied and incubated for 60 seconds. 3 microliters of solution was removed, then an additional 3 microliters of GFP-E892-Vt applied. After 60 seconds, 3 microliters of solution was removed, then the grid was blotted for 2 seconds before plunging.

-
Electron microscopy

MicroscopeFEI TECNAI 20
Electron beamAcceleration voltage: 120 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 137615 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 100000
Sample stageSpecimen holder model: GATAN LIQUID NITROGEN
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification.
DateJul 9, 2014
Image recordingCategory: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 212 / Average electron dose: 25 e/Å2

-
Image processing

CTF correctionDetails: FREALIGN (per segment)
Final reconstructionApplied symmetry - Helical parameters - Δz: 27.82 Å
Applied symmetry - Helical parameters - Δ&Phi: 166.77 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 16.4 Å / Resolution method: OTHER / Software - Name: CTFFIND3, EMAN2/SPARX, FREALIGN
DetailsMulti-model IHRSR was performed using EMAN2 / SPARX to select for bound segments, followed by single-model IHRSR with EMAN2 / SPARX and final reconstruction with FREALIGN (fixed helical parameters).

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more