[English] 日本語
Yorodumi
- PDB-8oi8: Cryo-EM structure of ADP-bound, filamentous beta-actin harboring ... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 8oi8
TitleCryo-EM structure of ADP-bound, filamentous beta-actin harboring the R183W mutation
ComponentsActin, cytoplasmic 1
KeywordsSTRUCTURAL PROTEIN / Actin filament / cytoskeletal protein / ATPase
Function / homology
Function and homology information


positive regulation of norepinephrine uptake / Regulation of CDH1 Function / Formation of the polybromo-BAF (pBAF) complex / Formation of the non-canonical BAF (ncBAF) complex / Formation of the canonical BAF (cBAF) complex / Formation of the embryonic stem cell BAF (esBAF) complex / Formation of neuronal progenitor and neuronal BAF (npBAF and nBAF) / bBAF complex / cellular response to cytochalasin B / npBAF complex ...positive regulation of norepinephrine uptake / Regulation of CDH1 Function / Formation of the polybromo-BAF (pBAF) complex / Formation of the non-canonical BAF (ncBAF) complex / Formation of the canonical BAF (cBAF) complex / Formation of the embryonic stem cell BAF (esBAF) complex / Formation of neuronal progenitor and neuronal BAF (npBAF and nBAF) / bBAF complex / cellular response to cytochalasin B / npBAF complex / nBAF complex / brahma complex / regulation of transepithelial transport / morphogenesis of a polarized epithelium / Formation of annular gap junctions / Formation of the dystrophin-glycoprotein complex (DGC) / structural constituent of postsynaptic actin cytoskeleton / GBAF complex / Gap junction degradation / Folding of actin by CCT/TriC / regulation of G0 to G1 transition / Cell-extracellular matrix interactions / protein localization to adherens junction / dense body / Tat protein binding / postsynaptic actin cytoskeleton / Prefoldin mediated transfer of substrate to CCT/TriC / RSC-type complex / regulation of double-strand break repair / regulation of nucleotide-excision repair / Adherens junctions interactions / RHOF GTPase cycle / adherens junction assembly / apical protein localization / Sensory processing of sound by inner hair cells of the cochlea / Sensory processing of sound by outer hair cells of the cochlea / Interaction between L1 and Ankyrins / tight junction / SWI/SNF complex / regulation of mitotic metaphase/anaphase transition / positive regulation of T cell differentiation / apical junction complex / positive regulation of double-strand break repair / maintenance of blood-brain barrier / regulation of norepinephrine uptake / nitric-oxide synthase binding / transporter regulator activity / cortical cytoskeleton / positive regulation of stem cell population maintenance / establishment or maintenance of cell polarity / NuA4 histone acetyltransferase complex / Recycling pathway of L1 / Regulation of MITF-M-dependent genes involved in pigmentation / brush border / regulation of G1/S transition of mitotic cell cycle / EPH-ephrin mediated repulsion of cells / negative regulation of cell differentiation / kinesin binding / RHO GTPases Activate WASPs and WAVEs / regulation of synaptic vesicle endocytosis / positive regulation of myoblast differentiation / RHO GTPases activate IQGAPs / regulation of protein localization to plasma membrane / positive regulation of double-strand break repair via homologous recombination / EPHB-mediated forward signaling / cytoskeleton organization / substantia nigra development / axonogenesis / calyx of Held / nitric-oxide synthase regulator activity / FCGR3A-mediated phagocytosis / actin filament / adherens junction / Translocation of SLC2A4 (GLUT4) to the plasma membrane / positive regulation of cell differentiation / cell motility / Regulation of endogenous retroelements by Piwi-interacting RNAs (piRNAs) / RHO GTPases Activate Formins / Signaling by high-kinase activity BRAF mutants / MAP2K and MAPK activation / Regulation of actin dynamics for phagocytic cup formation / kinetochore / structural constituent of cytoskeleton / B-WICH complex positively regulates rRNA expression / DNA Damage Recognition in GG-NER / VEGFA-VEGFR2 Pathway / platelet aggregation / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / tau protein binding / Schaffer collateral - CA1 synapse / nuclear matrix / cytoplasmic ribonucleoprotein granule / 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 / UCH proteinases / nucleosome
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
ADENOSINE-5'-DIPHOSPHATE / Actin, cytoplasmic 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.28 Å
AuthorsOosterheert, W. / Blanc, F.E.C. / Roy, A. / Belyy, A. / Hofnagel, O. / Hummer, G. / Bieling, P. / Raunser, S.
Funding support Germany, European Union, 3items
OrganizationGrant numberCountry
Max Planck Society Germany
Alexander von Humboldt Foundation Germany
European Research Council (ERC)856118European Union
CitationJournal: Nat Struct Mol Biol / Year: 2023
Title: Molecular mechanisms of inorganic-phosphate release from the core and barbed end of actin filaments.
Authors: Wout Oosterheert / Florian E C Blanc / Ankit Roy / Alexander Belyy / Micaela Boiero Sanders / Oliver Hofnagel / Gerhard Hummer / Peter Bieling / Stefan Raunser /
Abstract: The release of inorganic phosphate (P) from actin filaments constitutes a key step in their regulated turnover, which is fundamental to many cellular functions. The mechanisms underlying P release ...The release of inorganic phosphate (P) from actin filaments constitutes a key step in their regulated turnover, which is fundamental to many cellular functions. The mechanisms underlying P release from the core and barbed end of actin filaments remain unclear. Here, using human and bovine actin isoforms, we combine cryo-EM with molecular-dynamics simulations and in vitro reconstitution to demonstrate how actin releases P through a 'molecular backdoor'. While constantly open at the barbed end, the backdoor is predominantly closed in filament-core subunits and opens only transiently through concerted amino acid rearrangements. This explains why P escapes rapidly from the filament end but slowly from internal subunits. In a nemaline-myopathy-associated actin variant, the backdoor is predominantly open in filament-core subunits, resulting in accelerated P release and filaments with drastically shortened ADP-P caps. Our results provide the molecular basis for P release from actin and exemplify how a disease-linked mutation distorts the nucleotide-state distribution and atomic structure of the filament.
History
DepositionMar 22, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 16, 2023Provider: repository / Type: Initial release
Revision 1.1Sep 27, 2023Group: Database references / Refinement description / Category: citation / citation_author / em_3d_fitting_list
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.year / _em_3d_fitting_list.initial_refinement_model_id
Revision 1.2Oct 4, 2023Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.3Nov 22, 2023Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID

-
Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
C: Actin, cytoplasmic 1
A: Actin, cytoplasmic 1
B: Actin, cytoplasmic 1
D: Actin, cytoplasmic 1
E: Actin, cytoplasmic 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)211,22115
Polymers208,9635
Non-polymers2,25810
Water10,539585
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "C"
d_2ens_1chain "B"
d_3ens_1chain "A"
d_4ens_1chain "D"
d_5ens_1chain "E"

NCS domain segments:

Ens-ID: ens_1

Dom-IDComponent-IDBeg auth comp-IDBeg label comp-IDEnd auth comp-IDEnd label comp-IDAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
d_11ALAALAHISHISCA6 - 3716 - 371
d_12ADPADPADPADPCF401
d_21ALAALAHISHISBC6 - 3716 - 371
d_22ADPADPADPADPBJ401
d_31ALAALAHISHISAB6 - 3716 - 371
d_32ADPADPADPADPAH401
d_41ALAALAHISHISDD6 - 3716 - 371
d_42ADPADPADPADPDL401
d_51ALAALAHISHISEE6 - 3716 - 371
d_52ADPADPADPADPEN401

NCS oper:
IDCodeMatrixVector
1given(-0.964520151739, -0.233399926349, -0.123391050205), (0.226007438309, -0.971531230945, 0.0710472035138), (-0.13646067096, 0.0406391643467, 0.989811569746)309.858724101, 220.978695243, -13.2641646899
2given(0.892907583648, 0.449630210801, -0.0234247859805), (-0.450140015426, 0.892596618944, -0.0254016212381), (0.00948754845373, 0.0332257337631, 0.99940284022)-38.4865152081, 75.9709058504, -60.5148201639
3given(-0.962856689403, 0.232690506801, -0.136974901776), (-0.240307899472, -0.969798299055, 0.0417537136155), (-0.123122333974, 0.0731189933883, 0.989694146533)246.026188367, 288.064791549, 35.6738238829
4given(0.888893214479, -0.458113375947, 0.00099399763663), (0.457857439096, 0.888465099373, 0.0315647376143), (-0.0153433607184, -0.0276025718698, 0.999501215261)71.5248099114, -48.6543053678, 60.8597865492

-
Components

#1: Protein
Actin, cytoplasmic 1 / Beta-actin


Mass: 41792.633 Da / Num. of mol.: 5 / Mutation: R183W, C272A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ACTB / Plasmid: p2920 pFL_ACTB_R183W_C272A / Cell line (production host): BTI-Tnao38 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P60709, Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement
#2: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: ADP, energy-carrying molecule*YM
#3: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 585 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

-
Experimental details

-
Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: FILAMENT / 3D reconstruction method: single particle reconstruction

-
Sample preparation

ComponentName: Actin filament harboring the R183W mutation. / Type: COMPLEX
Details: Actin was purified as monomer from insect cells. It was then polymerized into a filament in vitro.
Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper) / Cell: BTI-Tnao38
Buffer solutionpH: 7.1
Details: 1x KMEH (10 mM HEPES pH 7.1, 100 mM KCl, 2 mM MgCl2, 1 mM EGTA) supplemented with 0.02% Tween20 (v/v)
Buffer component
IDConc.NameFormulaBuffer-ID
110 mMHEPES1
2100 mMpotassium chlorideKCl1
32 mMmagnesium chlorideMgCl21
41 mMEGTA1
50.02 % (v/v)Tween201
SpecimenConc.: 2.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Actin filaments were reconstituted by adding salt to monomeric actin.
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 286 K

-
Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Details: Titan Krios G3 microscope was aligned using Sherpa (FEI). Data collected in superresolution mode.
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: ZEMLIN TABLEAU
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 70 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 7916
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 15 eV

-
Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.20.1_4487refinement
PHENIX1.20.1_4487refinement
EM software
IDNameVersionCategory
1crYOLO1.5.8particle selection
2EPUimage acquisition
4CTFFIND4.13CTF correction
7Coot0.9.8.1model fitting
9PHENIX1.20.1-4487-000model refinement
10SPHIRE1.4initial Euler assignment
11RELION3.1.0final Euler assignment
12RELION3.1.0classification
13RELION3.1.03D reconstruction
Image processingDetails: K3 operated in super-resolution mode.
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1569882
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.28 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 1386604 / Details: Refinement from Local searches in RELION. / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Details: The structure was refined using phenix-real-space refine.
Atomic model buildingAccession code: 8a2t / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 31.83 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.003814630
ELECTRON MICROSCOPYf_angle_d0.539419870
ELECTRON MICROSCOPYf_chiral_restr0.04452215
ELECTRON MICROSCOPYf_plane_restr0.00352525
ELECTRON MICROSCOPYf_dihedral_angle_d11.47355395
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDRefine-IDTypeRms dev position (Å)
ens_1d_2AELECTRON MICROSCOPYNCS constraints2.34445256493E-11
ens_1d_3AELECTRON MICROSCOPYNCS constraints3.06042325467E-12
ens_1d_4AELECTRON MICROSCOPYNCS constraints8.247743843E-14
ens_1d_5AELECTRON MICROSCOPYNCS constraints2.40769520147E-12

+
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