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- PDB-9b52: RhAAV4282 Empty Capsid -

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Basic information

Entry
Database: PDB / ID: 9b52
TitleRhAAV4282 Empty Capsid
ComponentsCapsid protein VP1
KeywordsVIRAL PROTEIN / AAV / Adeno-Associated Virus / Empty Capsid
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 VP1
Function and homology information
Biological speciesAdeno-associated virus
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.57 Å
AuthorsDagotto, G. / Jenni, S. / Li, Z. / Barouch, D.H.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/Office of the DirectorOD011170 United States
CitationJournal: Mol Ther Methods Clin Dev / Year: 2024
Title: Identification of a novel neutralization epitope in rhesus AAVs.
Authors: Gabriel Dagotto / Jana L Fisher / David Li / Zhenyu Li / Simon Jenni / Zongli Li / Lawrence J Tartaglia / Peter Abbink / Dan H Barouch /
Abstract: Adeno-associated viruses (AAVs) are popular gene therapy delivery vectors, but their application can be limited by anti-vector immunity. Both preexisting neutralizing antibodies (NAbs) and post- ...Adeno-associated viruses (AAVs) are popular gene therapy delivery vectors, but their application can be limited by anti-vector immunity. Both preexisting neutralizing antibodies (NAbs) and post-administration NAbs can limit transgene expression and reduce the clinical utility of AAVs. The development of novel AAVs will advance our understanding of AAV immunity and may also have practical applications. In this study, we identified five novel AAV capsids from rhesus macaques. RhAAV4282 exhibited 91.4% capsid sequence similarity with AAV7 and showed similar tissue tropism with slightly diminished overall signal. Despite this sequence homology, RhAAV4282 and AAV7 showed limited cross-neutralization. We determined a cryo-EM structure of the RhAAV4282 capsid at 2.57 Å resolution and identified a small segment within the hypervariable region IV, involving seven amino acids that formed a shortened external loop in RhAAV4282 compared with AAV7. We generated RhAAV4282 and AAV7 mutants that involved swaps of this region and showed that this region partially determined neutralization phenotype. We termed this region the hypervariable region IV neutralizing epitope (HRNE). Our data suggests that modification of the HRNE can lead to AAVs with altered neutralization profiles.
History
DepositionMar 22, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 13, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
22: Capsid protein VP1


Theoretical massNumber of molelcules
Total (without water)58,1371
Polymers58,1371
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
SymmetryPoint symmetry: (Schoenflies symbol: I60 (icosahedral))

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Components

#1: Protein Capsid protein VP1


Mass: 58137.156 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Adeno-associated virus / Production host: Homo sapiens (human) / References: UniProt: B4Y879
Has protein modificationN

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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: Recombinant viral particles were grown using standard triple transfection method.
Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Adeno-associated virus
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293T
Details of virusEmpty: YES / Enveloped: NO / Isolate: STRAIN / Type: VIRUS-LIKE PARTICLE
Natural hostOrganism: Macaca mulatta
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company
MicroscopyModel: FEI POLARA 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1200 nm
Image recordingElectron dose: 52.5 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.57 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 35328 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 11.79 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00234233
ELECTRON MICROSCOPYf_angle_d0.46595777
ELECTRON MICROSCOPYf_chiral_restr0.0396605
ELECTRON MICROSCOPYf_plane_restr0.0031767
ELECTRON MICROSCOPYf_dihedral_angle_d5.343559

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