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- PDB-7ti4: Adeno-associated Virus Go.1 at 2.9 Angstroms resolution, AAVGo.1 ... -

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Basic information

Entry
Database: PDB / ID: 7ti4
TitleAdeno-associated Virus Go.1 at 2.9 Angstroms resolution, AAVGo.1 AAV-Go
ComponentsCapsid proteinCapsid
KeywordsVIRUS LIKE PARTICLE / AAVGo.1 / AAV / AAV-Go / AAVR / adeno-associated virus / PKD domain / parvovirus / virus / gene therapy / receptor / adeno-associated virus receptor / PKD1 / PKD
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 protein
Function and homology information
Biological speciesAdeno-associated virus
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.93 Å
AuthorsSilveria, M. / Large, E.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM122564 United States
CitationJournal: J Virol / Year: 2022
Title: Cross-Species Permissivity: Structure of a Goat Adeno-Associated Virus and Its Complex with the Human Receptor AAVR.
Authors: Edward E Large / Mark A Silveria / Onellah Weerakoon / Tommi A White / Michael S Chapman /
Abstract: Adeno-associated virus (AAV) is a small ssDNA satellite virus of high interest (in recombinant form) as a safe and effective gene therapy vector. AAV's human cell entry receptor (AAVR) contains ...Adeno-associated virus (AAV) is a small ssDNA satellite virus of high interest (in recombinant form) as a safe and effective gene therapy vector. AAV's human cell entry receptor (AAVR) contains polycystic kidney disease (PKD) domains bound by AAV. Seeking understanding of the spectrum of interactions, goat AAVGo.1 is investigated, because its host is the species most distant from human with reciprocal cross-species cell susceptibility. The structure of AAVGo.1, solved by cryo-EM to 2.9 Å resolution, is most similar to AAV5. Through ELISA (enzyme-linked immunosorbent assay) studies, it is shown that AAVGo.1 binds to human AAVR more strongly than do AAV2 or AAV5, and that it joins AAV5 in a class that binds exclusively to PKD domain 1 (PKD1), in contrast to other AAVs that interact primarily with PKD2. The AAVGo.1 cryo-EM structure of a complex with a PKD12 fragment of AAVR at 2.4 Å resolution shows PKD1 bound with minimal change in virus structure. There are only minor conformational adaptations in AAVR, but there is a near-rigid rotation of PKD1 with maximal displacement of the receptor domain by ~1 Å compared to PKD1 bound to AAV5. AAVGo.1 joins AAV5 as the second member of an emerging class of AAVs whose mode of receptor-binding is completely different from other AAVs, typified by AAV2. Adeno-associated virus (AAV) is a small ssDNA satellite parvovirus. As a recombinant vector with a protein shell encapsidating a transgene, recombinant AAV (rAAV) is a leading delivery vehicle for gene therapy, with two FDA-approved treatments and 150 clinical trials for 30 diseases. The human entry receptor AAVR has five PKD domains. To date, all serotypes, except AAV5, have interacted primarily with the second PKD domain, PKD2. Goat is the AAV host most distant from human with cross-species cell infectivity. AAVGo.1 is similar in structure to AAV5, the two forming a class with a distinct mode of receptor-binding. Within the two classes, binding interactions are mostly conserved, giving an indication of the latitude available in modulating delivery vectors.
History
DepositionJan 12, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 23, 2022Provider: repository / Type: Initial release
Revision 1.1Dec 14, 2022Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Jan 4, 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
Revision 1.3Jan 11, 2023Group: Database references / Category: citation / citation_author
Item: _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.4Jun 5, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Capsid protein


Theoretical massNumber of molelcules
Total (without water)80,6921
Polymers80,6921
Non-polymers00
Water0
1
A: Capsid protein
x 60


  • complete icosahedral assembly
  • Evidence: microscopy, Negative Stain EM and ELISA
  • 4.84 MDa, 60 polymers
Theoretical massNumber of molelcules
Total (without water)4,841,49660
Polymers4,841,49660
Non-polymers00
Water0
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 protein
x 5


  • icosahedral pentamer
  • 403 kDa, 5 polymers
Theoretical massNumber of molelcules
Total (without water)403,4585
Polymers403,4585
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation4
4
A: Capsid protein
x 6


  • icosahedral 23 hexamer
  • 484 kDa, 6 polymers
Theoretical massNumber of molelcules
Total (without water)484,1506
Polymers484,1506
Non-polymers00
Water0
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 protein / Capsid / VP1


Mass: 80691.594 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Adeno-associated virus / Gene: cap / Variant: Go.1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q5XXZ6

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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
Details: Expressed using SF9 cells with a pfastbac LIC vector. Purified with cesium chloride ultracentrifugation.
Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Adeno-associated virus / Strain: Go.1
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Details of virusEmpty: YES / Enveloped: NO / Isolate: SEROTYPE / Type: VIRUS-LIKE PARTICLE
Natural hostOrganism: Homo sapiens
Virus shellName: Capsid, VP3 / Diameter: 250 nm
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMHEPES1
225 mMMagnesium chlorideMgCl21
325 mMSodium chlorideNaClSodium chloride1
SpecimenConc.: 0.75 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Monodisperse
Specimen supportGrid material: COPPER / Grid type: PELCO Ultrathin Carbon with Lacey Carbon
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 293 K
Details: Two 2uL aliquots applied to grid (manual blotting between), prior to automated 3 second blot before plunging.

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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 FIELDBright-field microscopy / Nominal magnification: 64000 X / Nominal defocus max: 2700 nm / Nominal defocus min: 700 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 93 K / Temperature (min): 93 K
Image recordingElectron dose: 32.9 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)
Image scansWidth: 11520 / Height: 8184

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Processing

EM software
IDNameVersionCategory
10RELION3.1initial Euler assignment
13RELION3.13D reconstruction
CTF correctionDetails: CTF correction was performed per particle / Type: PHASE FLIPPING ONLY
Particle selectionDetails: LoG Picker was used for initial automated particle selection. Templates were then generated by 2D classification, followed by particle template selection in Relion 3.0.
3D reconstructionResolution: 2.93 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 140568 / Symmetry type: POINT

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