[English] 日本語
Yorodumi
- PDB-8of8: Cryo-EM structure of actomyosin-5a-S1 with the full-length lever ... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 8of8
TitleCryo-EM structure of actomyosin-5a-S1 with the full-length lever (nucleotide free)
Components
  • Actin, alpha skeletal muscle
  • Calmodulin-1
  • Unconventional myosin-Va
KeywordsMOTOR PROTEIN / myosin / cytoskeletal motor / myosin-va / myo5a / myosin-5a / S1 / rigor / nucleotide free / apo / lever / 6IQ / actomyosin / actomyosin-5a / actin / actomyosin-va / actin bound
Function / homology
Function and homology information


establishment of endoplasmic reticulum localization to postsynapse / regulation of postsynaptic cytosolic calcium ion concentration / CaMK IV-mediated phosphorylation of CREB / Cam-PDE 1 activation / CREB1 phosphorylation through the activation of CaMKII/CaMKK/CaMKIV cascasde / Glycogen breakdown (glycogenolysis) / Activation of RAC1 downstream of NMDARs / melanosome localization / endoplasmic reticulum localization / Reduction of cytosolic Ca++ levels ...establishment of endoplasmic reticulum localization to postsynapse / regulation of postsynaptic cytosolic calcium ion concentration / CaMK IV-mediated phosphorylation of CREB / Cam-PDE 1 activation / CREB1 phosphorylation through the activation of CaMKII/CaMKK/CaMKIV cascasde / Glycogen breakdown (glycogenolysis) / Activation of RAC1 downstream of NMDARs / melanosome localization / endoplasmic reticulum localization / Reduction of cytosolic Ca++ levels / Sodium/Calcium exchangers / Activation of Ca-permeable Kainate Receptor / Synthesis of IP3 and IP4 in the cytosol / CLEC7A (Dectin-1) induces NFAT activation / RHO GTPases activate PAKs / Calmodulin induced events / Inactivation, recovery and regulation of the phototransduction cascade / Tetrahydrobiopterin (BH4) synthesis, recycling, salvage and regulation / eNOS activation / Ion transport by P-type ATPases / Calcineurin activates NFAT / Unblocking of NMDA receptors, glutamate binding and activation / locomotion involved in locomotory behavior / Protein methylation / melanin metabolic process / RAF activation / VEGFR2 mediated vascular permeability / vesicle transport along actin filament / RAS processing / unconventional myosin complex / insulin-responsive compartment / FCERI mediated Ca+2 mobilization / Ca2+ pathway / RHO GTPases activate IQGAPs / Extra-nuclear estrogen signaling / RAF/MAP kinase cascade / PKA activation / developmental pigmentation / melanosome transport / secretory granule localization / Smooth Muscle Contraction / Regulation of MITF-M-dependent genes involved in pigmentation / Regulation of actin dynamics for phagocytic cup formation / actin filament-based movement / melanin biosynthetic process / hair follicle maturation / Platelet degranulation / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / melanocyte differentiation / actomyosin / Stimuli-sensing channels / Ion homeostasis / long-chain fatty acid biosynthetic process / myosin complex / insulin secretion / odontogenesis / intermediate filament / cytoskeletal motor activator activity / organelle localization by membrane tethering / pigmentation / mitochondrion-endoplasmic reticulum membrane tethering / autophagosome membrane docking / microfilament motor activity / myosin heavy chain binding / presynaptic endocytosis / regulation of cardiac muscle cell action potential / tropomyosin binding / negative regulation of ryanodine-sensitive calcium-release channel activity / troponin I binding / filamentous actin / exocytosis / calcineurin-mediated signaling / mesenchyme migration / actin filament bundle / cytoskeletal motor activity / actin filament bundle assembly / skeletal muscle myofibril / striated muscle thin filament / protein phosphatase activator activity / adenylate cyclase binding / regulation of ryanodine-sensitive calcium-release channel activity / skeletal muscle thin filament assembly / actin monomer binding / catalytic complex / photoreceptor outer segment / detection of calcium ion / regulation of cardiac muscle contraction / smooth endoplasmic reticulum / calcium channel inhibitor activity / cellular response to interferon-beta / stress fiber / skeletal muscle fiber development / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / vesicle-mediated transport / titin binding / voltage-gated potassium channel complex / actin filament polymerization / sperm midpiece / myelination / calcium channel complex
Similarity search - Function
Myosin 5a, cargo-binding domain / Class V myosin, motor domain / Dilute domain / DIL domain / Dilute domain profile. / DIL / IQ calmodulin-binding motif / Myosin, N-terminal, SH3-like / Myosin N-terminal SH3-like domain profile. / Short calmodulin-binding motif containing conserved Ile and Gln residues. ...Myosin 5a, cargo-binding domain / Class V myosin, motor domain / Dilute domain / DIL domain / Dilute domain profile. / DIL / IQ calmodulin-binding motif / Myosin, N-terminal, SH3-like / Myosin N-terminal SH3-like domain profile. / Short calmodulin-binding motif containing conserved Ile and Gln residues. / IQ motif, EF-hand binding site / Myosin head, motor domain / Myosin head (motor domain) / Myosin motor domain profile. / Myosin. Large ATPases. / IQ motif profile. / Kinesin motor domain 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 / EF-hand domain pair / EF-hand, calcium binding motif / ATPase, nucleotide binding domain / EF-Hand 1, calcium-binding site / EF-hand calcium-binding domain. / EF-hand calcium-binding domain profile. / EF-hand domain / EF-hand domain pair / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Calmodulin-1 / Actin, alpha skeletal muscle / Unconventional myosin-Va
Similarity search - Component
Biological speciesMus musculus (house mouse)
Oryctolagus cuniculus (rabbit)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 7.5 Å
AuthorsGravett, M.S.C. / Klebl, D.P. / Harlen, O.G. / Read, D.J. / Harris, S.A. / Muench, S.P. / Peckham, M.
Funding support United Kingdom, 5items
OrganizationGrant numberCountry
Wellcome Trust102174/B/13/Z United Kingdom
Wellcome Trust223125/Z/21/Z United Kingdom
Wellcome Trust108466/Z/15/Z United Kingdom
Engineering and Physical Sciences Research CouncilEP/T022167/1 United Kingdom
Engineering and Physical Sciences Research CouncilEP/R013012 United Kingdom
CitationJournal: Structure / Year: 2024
Title: Exploiting cryo-EM structures of actomyosin-5a to reveal the physical properties of its lever.
Authors: Molly S C Gravett / David P Klebl / Oliver G Harlen / Daniel J Read / Stephen P Muench / Sarah A Harris / Michelle Peckham /
Abstract: Myosin 5a (Myo5a) is a dimeric processive motor protein that transports cellular cargos along filamentous actin (F-actin). Its long lever is responsible for its large power-stroke, step size, and ...Myosin 5a (Myo5a) is a dimeric processive motor protein that transports cellular cargos along filamentous actin (F-actin). Its long lever is responsible for its large power-stroke, step size, and load-bearing ability. Little is known about the levers' structure and physical properties, and how they contribute to walking mechanics. Using cryoelectron microscopy (cryo-EM) and molecular dynamics (MD) simulations, we resolved the structure of monomeric Myo5a, comprising the motor domain and full-length lever, bound to F-actin. The range of its lever conformations revealed its physical properties, how stiffness varies along its length and predicts a large, 35 nm, working stroke. Thus, the newly released trail head in a dimeric Myo5a would only need to perform a small diffusive search for its new binding site on F-actin, and stress would only be generated across the dimer once phosphate is released from the lead head, revealing new insight into the walking behavior of Myo5a.
History
DepositionMar 14, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Sep 25, 2024Provider: repository / Type: Initial release
Revision 1.1Apr 9, 2025Group: Data collection / Database references / Structure summary
Category: citation / citation_author ...citation / citation_author / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name / _em_admin.last_update / _pdbx_entry_details.has_protein_modification

-
Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: Calmodulin-1
B: Calmodulin-1
C: Calmodulin-1
D: Calmodulin-1
E: Calmodulin-1
F: Calmodulin-1
J: Actin, alpha skeletal muscle
K: Actin, alpha skeletal muscle
L: Actin, alpha skeletal muscle
M: Unconventional myosin-Va


Theoretical massNumber of molelcules
Total (without water)332,55110
Polymers332,55110
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

-
Components

#1: Protein
Calmodulin-1


Mass: 16721.350 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Calm1, Calm, Cam, Cam1 / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P0DP26
#2: Protein Actin, alpha skeletal muscle / Alpha-actin-1


Mass: 41875.633 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Details: HIC is tele-methylhistidine / Source: (natural) Oryctolagus cuniculus (rabbit) / Tissue: skeletal muscle / References: UniProt: P68135
#3: Protein Unconventional myosin-Va / Dilute myosin heavy chain / non-muscle


Mass: 106596.195 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Myo5a, Dilute / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q99104
Has ligand of interestN
Has protein modificationY

-
Experimental details

-
Experiment

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

-
Sample preparation

Component
IDNameTypeEntity IDParent-IDSourceDetails
1Myosin-5a bound to F-actinCOMPLEXall0MULTIPLE SOURCES
2Actin, alpha skeletal muscleCOMPLEX#21NATURAL
3Unconventional myosin-Va with calmodulin-1 boundCOMPLEX#31RECOMBINANT
4Calmodulin-1COMPLEX#11RECOMBINANT1 calmodulin bound per IQ domain (6 overall)
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
110.33 MDaNO
2115 kDa/nmNO
310.11 MDaNO
510.02 MDaNO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-IDOrgan
22Oryctolagus cuniculus (rabbit)9986skeletal muscle
33Mus musculus (house mouse)10090
54Mus musculus (house mouse)10090
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
33Spodoptera frugiperda (fall armyworm)7108
54Spodoptera frugiperda (fall armyworm)7108
Buffer solutionpH: 7
Buffer component
IDConc.NameFormulaBuffer-ID
1100 mMpotassium chlorideKCl1
20.1 mMegtazic acid (EGTA)C14H24N2O101
31 mMmagnesium chlorideMgCl21
410 mM3-morpholinopropane-1-sulfonic acid (MOPS)C7H15NO4S1
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: 5.26 mg/mL Actin, alpha skeletal muscle 266 mg/mL FlAG-tagged unconventional myosin-Va (residues 1-907) 84.2 mg/mL Calmodulin-1
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 80 % / Chamber temperature: 281.15 K

-
Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 75000 X / Nominal defocus max: 3600 nm / Nominal defocus min: 1800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 1.5 sec. / Electron dose: 63.13 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4285

-
Processing

EM software
IDNameVersionCategoryDetails
1RELION3.1particle selectionmanual picking helical start end coordinates
4Gctf1.18CTF correction
7ISOLDEmodel fitting
8Ambermodel fitting
11RELION3.1initial Euler assignment
12RELION3.1final Euler assignment
13RELION3.1classification
14RELION3.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 838213
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 7.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 22337 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model building

3D fitting-ID: 1

IDPDB-IDAccession codeChain-IDDetailsInitial refinement model-IDSource nameType
17PLUDchain D was used as a template to model the motor domain1SwissModelin silico model
2the lever structure with CaM bound was predicted in AlphaFoldAlphaFoldin silico model
37PLU

7plu

7PLUchains J-L were used to model actin3PDBexperimental model

+
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