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- PDB-7q8c: Leishmania major actin filament in ADP-state -

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Basic information

Entry
Database: PDB / ID: 7q8c
TitleLeishmania major actin filament in ADP-state
ComponentsActin
KeywordsSTRUCTURAL PROTEIN / Actin / Filament / Parasite / ADP-Pi
Function / homology
Function and homology information


kinetoplast / actin cytoskeleton / endonuclease activity / chromatin remodeling
Similarity search - Function
ATPase, substrate binding domain, subdomain 4 / Actin; Chain A, domain 4 / 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, substrate binding domain, subdomain 4 / Actin; Chain A, domain 4 / 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 / Alpha-Beta Complex / Alpha Beta
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / Actin
Similarity search - Component
Biological speciesLeishmania major (eukaryote)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 2.72 Å
AuthorsKotila, T. / Muniyandi, S. / Lappalainen, P. / Huiskonen, J.T.
Funding support Finland, 2items
OrganizationGrant numberCountry
Academy of Finland320161 Finland
Jane and Aatos Erkko Foundation4708679 Finland
CitationJournal: Nat Commun / Year: 2022
Title: Structural basis of rapid actin dynamics in the evolutionarily divergent Leishmania parasite.
Authors: Tommi Kotila / Hugo Wioland / Muniyandi Selvaraj / Konstantin Kogan / Lina Antenucci / Antoine Jégou / Juha T Huiskonen / Guillaume Romet-Lemonne / Pekka Lappalainen /
Abstract: Actin polymerization generates forces for cellular processes throughout the eukaryotic kingdom, but our understanding of the 'ancient' actin turnover machineries is limited. We show that, ...Actin polymerization generates forces for cellular processes throughout the eukaryotic kingdom, but our understanding of the 'ancient' actin turnover machineries is limited. We show that, despite > 1 billion years of evolution, pathogenic Leishmania major parasite and mammalian actins share the same overall fold and co-polymerize with each other. Interestingly, Leishmania harbors a simple actin-regulatory machinery that lacks cofilin 'cofactors', which accelerate filament disassembly in higher eukaryotes. By applying single-filament biochemistry we discovered that, compared to mammalian proteins, Leishmania actin filaments depolymerize more rapidly from both ends, and are severed > 100-fold more efficiently by cofilin. Our high-resolution cryo-EM structures of Leishmania ADP-, ADP-Pi- and cofilin-actin filaments identify specific features at actin subunit interfaces and cofilin-actin interactions that explain the unusually rapid dynamics of parasite actin filaments. Our findings reveal how divergent parasites achieve rapid actin dynamics using a remarkably simple set of actin-binding proteins, and elucidate evolution of the actin cytoskeleton.
History
DepositionNov 11, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 18, 2022Provider: repository / Type: Initial release
Revision 1.1Jun 29, 2022Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _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
Revision 1.2Jul 17, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_3d_fitting_list / em_admin / pdbx_initial_refinement_model
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Actin
B: Actin
C: Actin
D: Actin
E: Actin
hetero molecules


Theoretical massNumber of molelcules
Total (without water)212,57715
Polymers210,3195
Non-polymers2,25810
Water6,485360
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: assay for oligomerization
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area15080 Å2
ΔGint-141 kcal/mol
Surface area77770 Å2

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Components

#1: Protein
Actin


Mass: 42063.867 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Leishmania major (eukaryote) / Gene: ACT, LMJF_04_1230 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9U1E8
#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.: 360 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

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Experimental details

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Experiment

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

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Sample preparation

ComponentName: Bare, undecorated, ADP-state actin filament from the sample mixed with Leishmania major cofilin.
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightUnits: KILODALTONS/NANOMETER / Experimental value: NO
Source (natural)Organism: Leishmania major (eukaryote)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.4
Details: 10 mM HEPES, 125 mM NaCl, 5 mM KCl, 0.2 mM ATP, 0.4 mM EGTA, 1 mM MgCl2, 1 mM DTT
SpecimenConc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 279.15 K / Details: blot for 5 seconds before plunging, blot force 15

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 55 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of real images: 50

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameVersionCategoryDetails
1RELION3.1particle selection
4CTFFIND4.1CTF correction
7UCSF Chimera1.14model fittingbuild 42094
8Coot0.8.9.1model fitting
10RELION3.1initial Euler assignment
11RELION3.1final Euler assignment
12RELION3.1classification
13RELION3.13D reconstruction
14UCSF ChimeraX1.1.1model refinement
15ISOLDE1.1.0model refinement
16PHENIX1.19.2model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Helical symmertyAngular rotation/subunit: -166.54 ° / Axial rise/subunit: 28.4 Å / Axial symmetry: C1
3D reconstructionResolution: 2.72 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 232527 / Algorithm: FOURIER SPACE / Symmetry type: HELICAL
Atomic model buildingSpace: REAL
Atomic model buildingPDB-ID: 6DJO

6djo


Pdb chain-ID: A / Accession code: 6DJO / Source name: PDB / Type: experimental model

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