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- PDB-7thr: Cryo-electron microscopy of Adeno-associated virus serotype 4 at 2.2 A -

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Basic information

Entry
Database: PDB / ID: 7thr
TitleCryo-electron microscopy of Adeno-associated virus serotype 4 at 2.2 A
ComponentsCapsid
KeywordsVIRUS LIKE PARTICLE / AAV4 / adeno-associated virus / serotype 4
Function / homologyPhospholipase A2-like domain / Phospholipase A2-like domain / Parvovirus coat protein VP2 / Parvovirus coat protein VP1/VP2 / Parvovirus coat protein VP2 / Capsid/spike protein, ssDNA virus / T=1 icosahedral viral capsid / structural molecule activity / Capsid
Function and homology information
Biological speciesAdeno-associated virus - 4
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.21 Å
AuthorsZane, G.M. / Silveria, M.A. / Meyer, N.L. / White, T.A. / Chapman, M.S.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM122564 United States
CitationJournal: Acta Crystallogr D Struct Biol / Year: 2023
Title: Cryo-EM structure of adeno-associated virus 4 at 2.2 Å resolution.
Authors: Grant Zane / Mark Silveria / Nancy Meyer / Tommi White / Rui Duan / Xiaoqin Zou / Michael Chapman /
Abstract: Adeno-associated virus (AAV) is the vector of choice for several approved gene-therapy treatments and is the basis for many ongoing clinical trials. Various strains of AAV exist (referred to as ...Adeno-associated virus (AAV) is the vector of choice for several approved gene-therapy treatments and is the basis for many ongoing clinical trials. Various strains of AAV exist (referred to as serotypes), each with their own transfection characteristics. Here, a high-resolution cryo-electron microscopy structure (2.2 Å) of AAV serotype 4 (AAV4) is presented. The receptor responsible for transduction of the AAV4 clade of AAV viruses (including AAV11, AAV12 and AAVrh32.33) is unknown. Other AAVs interact with the same cell receptor, adeno-associated virus receptor (AAVR), in one of two different ways. AAV5-like viruses interact exclusively with the polycystic kidney disease-like 1 (PKD1) domain of AAVR, while most other AAVs interact primarily with the PKD2 domain. A comparison of the present AAV4 structure with prior corresponding structures of AAV5, AAV2 and AAV1 in complex with AAVR provides a foundation for understanding why the AAV4-like clade is unable to interact with either PKD1 or PKD2 of AAVR. The conformation of the AAV4 capsid in variable regions I, III, IV and V on the viral surface appears to be sufficiently different from AAV2 to ablate binding with PKD2. Differences between AAV4 and AAV5 in variable region VII appear to be sufficient to exclude binding with PKD1.
History
DepositionJan 11, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 25, 2023Provider: repository / Type: Initial release
Revision 1.1Aug 9, 2023Group: Database references / Category: citation / citation_author
Item: _citation.journal_abbrev / _citation.journal_id_CSD ..._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
Revision 1.2Jun 5, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / em_3d_fitting_list
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

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Capsid
hetero molecules


Theoretical massNumber of molelcules
Total (without water)80,7373
Polymers80,6881
Non-polymers492
Water3,945219
1
A: Capsid
hetero molecules
x 60


  • complete icosahedral assembly
  • Evidence: electron microscopy
  • 4.84 MDa, 60 polymers
Theoretical massNumber of molelcules
Total (without water)4,844,198180
Polymers4,841,28160
Non-polymers2,917120
Water1,08160
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation59
2


  • Idetical with deposited unit
  • icosahedral asymmetric unit
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
A: Capsid
hetero molecules
x 5


  • icosahedral pentamer
  • 404 kDa, 5 polymers
Theoretical massNumber of molelcules
Total (without water)403,68315
Polymers403,4405
Non-polymers24310
Water905
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation4
4
A: Capsid
hetero molecules
x 6


  • icosahedral 23 hexamer
  • 484 kDa, 6 polymers
Theoretical massNumber of molelcules
Total (without water)484,42018
Polymers484,1286
Non-polymers29212
Water1086
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation5
5


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit, std point frame
TypeNameSymmetry operationNumber
transform to point frame1
SymmetryPoint symmetry: (Schoenflies symbol: I (icosahedral))

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Components

#1: Protein Capsid


Mass: 80688.023 Da / Num. of mol.: 1 / Mutation: M1T
Source method: isolated from a genetically manipulated source
Details: VP1 sequence of AAV4 / Source: (gene. exp.) Adeno-associated virus - 4 / Plasmid: pFastBac-LIC(4A) / Details (production host): AmpR / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: O41855
#2: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 219 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN

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Experimental details

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Experiment

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

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Sample preparation

ComponentName: Adeno-associated virus / Type: VIRUS / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 3.746 MDa / Experimental value: NO
Source (natural)Organism: Adeno-associated virus / Strain: Serotype 4
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm) / Strain: Sf9 / Plasmid: pFastBac-LIC(4A)
Details of virusEmpty: YES / Enveloped: NO / Isolate: SEROTYPE / Type: VIRUS-LIKE PARTICLE
Natural hostOrganism: Homo sapiens
Virus shellName: Capsid / Diameter: 250 nm / Triangulation number (T number): 1
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
1150 mMsodium chlorideNaCl1
225 mMHEPESHEPES1
SpecimenConc.: 0.33 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: The sample was isolated by multiple rounds of cesium chloride density centrifugation and dialyzed into a HEPES-buffered, sodium chloride solution (see Meyer et al., 2019 Bioprotocols paper).
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: PELCO Ultrathin Carbon with Lacey Carbon
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 298 K / Details: blot force: 4 time: 2 s

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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 magnification: 16500 X / Nominal defocus max: 1800 nm / Nominal defocus min: 600 nm / C2 aperture diameter: 50 µm
Image recordingAverage exposure time: 1.4 sec. / Electron dose: 48 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4809
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategoryDetails
2SerialEM3.8image acquisitionAt PNCC
4RELION3.1.1CTF correction
7CNS0.5model fittingpackaged with PaStO
9Coot0.8.9.2model refinement
10RELION3.1.1initial Euler assignment
11RELION3.1.1final Euler assignment
12RELION3.1.1classification
13RELION3.1.13D reconstruction
CTF correctionType: PHASE FLIPPING ONLY
Particle selectionNum. of particles selected: 75226
SymmetryPoint symmetry: I (icosahedral)
3D reconstructionResolution: 2.21 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 75226 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingDetails: Link to CNS_RSRef software: https://chapman.missouri.edu/software/
Atomic model buildingPDB-ID: 2G8G

2g8g


Pdb chain-ID: A / Accession code: 2G8G / Pdb chain residue range: 211-734 / Source name: PDB / Type: experimental model

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