[English] 日本語
Yorodumi
- EMDB-13328: Time-resolved cryo-EM structures of ATP-induced actomyosin dissoc... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-13328
TitleTime-resolved cryo-EM structures of ATP-induced actomyosin dissociation
Map datasharpened map of actomyosin from all timepoints combined
Sample
  • Complex: Skeletal myosin subfragment 1 (A1 fraction) from rabbit in complex with filamentous actin from rabbit
Biological speciesOryctolagus cuniculus (rabbit)
Methodhelical reconstruction / cryo EM / Resolution: 7.5 Å
AuthorsKlebl DP / White HD / Sobott F / Muench SP
Funding support3 items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)BB/P0263971/1
American Heart AssociationAMR21-236078
Wellcome Trust08466/Z/15/Z
CitationJournal: Acta Crystallogr D Struct Biol / Year: 2021
Title: On-grid and in-flow mixing for time-resolved cryo-EM.
Authors: David P Klebl / Howard D White / Frank Sobott / Stephen P Muench /
Abstract: Time-resolved cryo-electron microscopy (TrEM) allows the study of proteins under non-equilibrium conditions on the millisecond timescale, permitting the analysis of large-scale conformational changes ...Time-resolved cryo-electron microscopy (TrEM) allows the study of proteins under non-equilibrium conditions on the millisecond timescale, permitting the analysis of large-scale conformational changes or assembly and disassembly processes. However, the technique is developing and there have been few comparisons with other biochemical kinetic studies. Using current methods, the shortest time delay is on the millisecond timescale (∼5-10 ms), given by the delay between sample application and vitrification, and generating longer time points requires additional approaches such as using a longer delay line between the mixing element and nozzle, or an incubation step on the grid. To compare approaches, the reaction of ATP with the skeletal actomyosin S1 complex was followed on grids prepared with a 7-700 ms delay between mixing and vitrification. Classification of the cryo-EM data allows kinetic information to be derived which agrees with previous biochemical measurements, showing fast dissociation, low occupancy during steady-state hydrolysis and rebinding once ATP has been hydrolysed. However, this rebinding effect is much less pronounced when on-grid mixing is used and may be influenced by interactions with the air-water interface. Moreover, in-flow mixing results in a broader distribution of reaction times due to the range of velocities in a laminar flow profile (temporal spread), especially for longer time delays. This work shows the potential of TrEM, but also highlights challenges and opportunities for further development.
History
DepositionAug 4, 2021-
Header (metadata) releaseOct 13, 2021-
Map releaseOct 13, 2021-
UpdateOct 13, 2021-
Current statusOct 13, 2021Processing site: PDBe / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.05
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.05
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_13328.png

Downloads & links

-
Map

FileDownload / File: emd_13328.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationsharpened map of actomyosin from all timepoints combined
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.13 Å/pix.
x 200 pix.
= 426. Å		2.13 Å/pix.
x 200 pix.
= 426. Å		2.13 Å/pix.
x 200 pix.
= 426. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.13 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05 / Movie #1: 0.05
Minimum - Maximum-0.043096405 - 0.14011763
Average (Standard dev.)0.00021942142 (±0.0042872727)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 426.00003 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.132.132.13
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z426.000426.000426.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.0430.1400.000

-
Supplemental data

+
Mask #1

Fileemd_13328_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: unsharpened reconstruction from all 13 ms particles

Fileemd_13328_additional_1.map
Annotationunsharpened reconstruction from all 13 ms particles
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: unsharpened reconstruction from all 340 ms particles

Fileemd_13328_additional_10.map
Annotationunsharpened reconstruction from all 340 ms particles
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: unsharpened reconstruction from all 400 ms particles

Fileemd_13328_additional_11.map
Annotationunsharpened reconstruction from all 400 ms particles
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: unsharpened reconstruction from all 640 ms particles

Fileemd_13328_additional_2.map
Annotationunsharpened reconstruction from all 640 ms particles
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: halfmap 2 of actin from all timepoints combined

Fileemd_13328_additional_3.map
Annotationhalfmap 2 of actin from all timepoints combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: unsharpened map of actomyosin from all timepoints combined

Fileemd_13328_additional_4.map
Annotationunsharpened map of actomyosin from all timepoints combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: sharpened map of actin from all timepoints combined

Fileemd_13328_additional_5.map
Annotationsharpened map of actin from all timepoints combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: unsharpened map of actin from all timepoints combined

Fileemd_13328_additional_6.map
Annotationunsharpened map of actin from all timepoints combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: halfmap 1 of actin from all timepoints combined

Fileemd_13328_additional_7.map
Annotationhalfmap 1 of actin from all timepoints combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: unsharpened reconstruction from all 700 ms particles

Fileemd_13328_additional_8.map
Annotationunsharpened reconstruction from all 700 ms particles
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Additional map: unsharpened reconstruction from all 7 ms particles

Fileemd_13328_additional_9.map
Annotationunsharpened reconstruction from all 7 ms particles
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Half map: halfmap 1 of actomyosin from all timepoints combined

Fileemd_13328_half_map_1.map
Annotationhalfmap 1 of actomyosin from all timepoints combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

+
Half map: halfmap 2 of actomyosin from all timepoints combined

Fileemd_13328_half_map_2.map
Annotationhalfmap 2 of actomyosin from all timepoints combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : Skeletal myosin subfragment 1 (A1 fraction) from rabbit in comple...

EntireName: Skeletal myosin subfragment 1 (A1 fraction) from rabbit in complex with filamentous actin from rabbit
Components
  • Complex: Skeletal myosin subfragment 1 (A1 fraction) from rabbit in complex with filamentous actin from rabbit

-
Supramolecule #1: Skeletal myosin subfragment 1 (A1 fraction) from rabbit in comple...

SupramoleculeName: Skeletal myosin subfragment 1 (A1 fraction) from rabbit in complex with filamentous actin from rabbit
type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Oryctolagus cuniculus (rabbit)

-
Experimental details

-
Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

-
Sample preparation

BufferpH: 7
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300
VitrificationCryogen name: ETHANE / Chamber temperature: 293 K / Instrument: HOMEMADE PLUNGER

-
Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: INTEGRATING / Number grids imaged: 6 / Average electron dose: 62.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

-
Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 27.56 Å
Applied symmetry - Helical parameters - Δ&Phi: -166.25 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 7.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 74556
Startup modelType of model: OTHER / Details: featureless cylinder
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

+
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