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- PDB-8cco: JAS-stabilized F-ActinII from Plasmodium falciparum -

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Basic information

Entry
Database: PDB / ID: 8cco
TitleJAS-stabilized F-ActinII from Plasmodium falciparum
ComponentsActin-2
KeywordsCELL INVASION / Filament-forming male-gametogenesis Zygote Hydrolase ATP-binding Jasplakinolide malaria methylation
Function / homology
Function and homology information


Platelet degranulation / symbiont-mediated actin polymerization-dependent cell-to-cell migration in host / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / Neutrophil degranulation / microfilament motor activity / cytoskeleton organization / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / structural constituent of cytoskeleton / hydrolase activity ...Platelet degranulation / symbiont-mediated actin polymerization-dependent cell-to-cell migration in host / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / Neutrophil degranulation / microfilament motor activity / cytoskeleton organization / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / structural constituent of cytoskeleton / hydrolase activity / ATP binding / cytoplasm
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
Jasplakinolide / ADENOSINE-5'-DIPHOSPHATE / Actin-2
Similarity search - Component
Biological speciesPlasmodium falciparum (malaria parasite P. falciparum)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 3.28 Å
AuthorsKursula, I. / Lopez, A.J.
Funding support Finland, Norway, United Kingdom, 5items
OrganizationGrant numberCountry
Academy of Finland Finland
Norwegian Research Council Norway
Sigrid Juselius Foundation Finland
Cancer Research UK United Kingdom
Medical Research Council (MRC, United Kingdom) United Kingdom
CitationJournal: PLoS Pathog / Year: 2023
Title: Structure and function of Plasmodium actin II in the parasite mosquito stages.
Authors: Andrea J Lopez / Maria Andreadaki / Juha Vahokoski / Elena Deligianni / Lesley J Calder / Serena Camerini / Anika Freitag / Ulrich Bergmann / Peter B Rosenthal / Inga Sidén-Kiamos / Inari Kursula /
Abstract: Actins are filament-forming, highly-conserved proteins in eukaryotes. They are involved in essential processes in the cytoplasm and also have nuclear functions. Malaria parasites (Plasmodium spp.) ...Actins are filament-forming, highly-conserved proteins in eukaryotes. They are involved in essential processes in the cytoplasm and also have nuclear functions. Malaria parasites (Plasmodium spp.) have two actin isoforms that differ from each other and from canonical actins in structure and filament-forming properties. Actin I has an essential role in motility and is fairly well characterized. The structure and function of actin II are not as well understood, but mutational analyses have revealed two essential functions in male gametogenesis and in the oocyst. Here, we present expression analysis, high-resolution filament structures, and biochemical characterization of Plasmodium actin II. We confirm expression in male gametocytes and zygotes and show that actin II is associated with the nucleus in both stages in filament-like structures. Unlike actin I, actin II readily forms long filaments in vitro, and near-atomic structures in the presence or absence of jasplakinolide reveal very similar structures. Small but significant differences compared to other actins in the openness and twist, the active site, the D-loop, and the plug region contribute to filament stability. The function of actin II was investigated through mutational analysis, suggesting that long and stable filaments are necessary for male gametogenesis, while a second function in the oocyst stage also requires fine-tuned regulation by methylation of histidine 73. Actin II polymerizes via the classical nucleation-elongation mechanism and has a critical concentration of ~0.1 μM at the steady-state, like actin I and canonical actins. Similarly to actin I, dimers are a stable form of actin II at equilibrium.
History
DepositionJan 27, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 1, 2023Provider: repository / Type: Initial release
Revision 1.1Mar 22, 2023Group: Database references / Category: citation / citation_author / Item: _citation_author.identifier_ORCID
Revision 1.2Aug 23, 2023Group: Data collection / Database references / Source and taxonomy
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / entity_src_gen / struct_ref / struct_ref_seq / struct_ref_seq_dif
Item: _entity_src_gen.pdbx_gene_src_gene / _struct_ref.db_code ..._entity_src_gen.pdbx_gene_src_gene / _struct_ref.db_code / _struct_ref.pdbx_db_accession / _struct_ref_seq.pdbx_db_accession / _struct_ref_seq_dif.pdbx_seq_db_accession_code

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Actin-2
B: Actin-2
C: Actin-2
D: Actin-2
E: Actin-2
F: Actin-2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)263,74824
Polymers256,7816
Non-polymers6,96718
Water00
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_1(chain "F" and (resid 5 through 94 or resid 96...
d_2ens_1(chain "B" and (resid 5 through 94 or resid 96...
d_3ens_1(chain "C" and (resid 5 through 94 or resid 96...
d_4ens_1(chain "D" and (resid 5 through 94 or resid 96...
d_5ens_1(chain "E" and (resid 5 through 94 or resid 96...
d_6ens_1(chain "A" and (resid 5 through 94 or resid 96...

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1ALALEUU1 - 90
d_12ens_1VALGLUU92 - 201
d_13ens_1GLUASPU203 - 220
d_14ens_1GLUARGU222 - 287
d_15ens_1GLUPHEU289 - 372
d_16ens_19UE9UEV
d_17ens_1ADPADPW
d_21ens_1ALALEUE1 - 90
d_22ens_1VALGLUE92 - 201
d_23ens_1GLUASPE203 - 220
d_24ens_1GLUARGE222 - 287
d_25ens_1GLUPHEE289 - 372
d_26ens_19UE9UEF
d_27ens_1ADPADPG
d_31ens_1ALALEUI1 - 90
d_32ens_1VALGLUI92 - 201
d_33ens_1GLUASPI203 - 220
d_34ens_1GLUARGI222 - 287
d_35ens_1GLUPHEI289 - 372
d_36ens_19UE9UEJ
d_37ens_1ADPADPK
d_41ens_1ALALEUM1 - 90
d_42ens_1VALGLUM92 - 201
d_43ens_1GLUASPM203 - 220
d_44ens_1GLUARGM222 - 287
d_45ens_1GLUPHEM289 - 372
d_46ens_19UE9UEN
d_47ens_1ADPADPO
d_51ens_1ALALEUQ1 - 90
d_52ens_1VALGLUQ92 - 201
d_53ens_1GLUASPQ203 - 220
d_54ens_1GLUARGQ222 - 287
d_55ens_1GLUPHEQ289 - 372
d_56ens_19UE9UER
d_57ens_1ADPADPS
d_61ens_1ALALEUA1 - 90
d_62ens_1VALGLUA92 - 201
d_63ens_1GLUASPA203 - 220
d_64ens_1GLUARGA222 - 287
d_65ens_1GLUPHEA289 - 372
d_66ens_19UE9UEB
d_67ens_1ADPADPC

NCS oper:
IDCodeMatrixVector
1given(0.611732634111, 0.791064573336, 0.000158673847152), (-0.791064581658, 0.611732588147, 0.000261238864927), (0.000109590848033, -0.000285329599491, 0.999999953288)-72.0532801907, 210.738819005, -113.423995308
2given(-0.774597307121, -0.632454703499, -0.000244587775842), (0.63245474285, -0.774597300761, -0.000141069555265), (-0.000100236927219, -0.0002639627965, 0.999999960138)430.362916984, 204.218069948, -85.0319268886
3given(0.897747473633, 0.440510460464, 8.8359606006E-5), (-0.440510468791, 0.897747444601, 0.000229342799432), (2.17032916838E-5, -0.00024481525025, 0.999999969797)-60.4979235864, 96.9592795412, -56.6902333853
4given(-0.97404055673, -0.226373505704, -0.000172514043283), (0.226373529584, -0.974040557212, -0.000134198509484), (-0.000137656687794, -0.00016976740377, 0.999999976115)393.397067711, 312.46092183, -28.3133033427
5given(-0.416855447759, -0.908972767481, -0.000208927308672), (0.908972782525, -0.416855400348, -0.00023628345675), (0.000127682750692, -0.00028840528327, 0.99999995026)415.830928878, 90.8645781628, -141.788532395

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Components

#1: Protein
Actin-2 / Actin II


Mass: 42796.879 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Plasmodium falciparum (malaria parasite P. falciparum)
Gene: ACT2, ACTII, PF14_0124, PF3D7_1412500 / Plasmid: pFastBacTHT A / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: Q8ILW9, Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement
#2: Chemical
ChemComp-9UE / Jasplakinolide


Mass: 709.670 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C36H45BrN4O6 / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: ADP, energy-carrying molecule*YM
#4: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
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: JAS-stabilized F-ActinII / Type: COMPLEX / Details: Actin in the ADP-Mg state / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Plasmodium falciparum (malaria parasite P. falciparum)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm) / Cell: Sf9 / Plasmid: pFastBacTHT A
Buffer solutionpH: 7.5
Details: 10 mM HEPES pH 7.5, 0.2 mM CaCl2, 0.5 mM ATP, 0.5 mM TCEP, 4 mM MgCl2, 50 mM KCl, 10 mM EGTA
SpecimenConc.: 0.43 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE

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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 / Nominal defocus max: 2600 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 47.2 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k) / Num. of real images: 3977

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameVersionCategory
1RELION3.1betaparticle selection
4CTFFIND4.1.10CTF correction
7UCSF Chimera1.14model fitting
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Helical symmertyAngular rotation/subunit: -166.9 ° / Axial rise/subunit: 28.37 Å / Axial symmetry: C1
Particle selectionNum. of particles selected: 448000
3D reconstructionResolution: 3.28 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 272000 / Symmetry type: HELICAL
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 38.83 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.001618762
ELECTRON MICROSCOPYf_angle_d0.485525470
ELECTRON MICROSCOPYf_chiral_restr0.04292802
ELECTRON MICROSCOPYf_plane_restr0.00343240
ELECTRON MICROSCOPYf_dihedral_angle_d9.88617086
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDRefine-IDTypeRms dev position (Å)
ens_1d_2EELECTRON MICROSCOPYNCS constraints0.000712442248243
ens_1d_3EELECTRON MICROSCOPYNCS constraints0.000713081990231
ens_1d_4EELECTRON MICROSCOPYNCS constraints0.000706957244935
ens_1d_5EELECTRON MICROSCOPYNCS constraints0.000713573191272
ens_1d_6EELECTRON MICROSCOPYNCS constraints0.000711512028116

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