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- EMDB-29377: AAV1 VP3 Only Capsid -

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Basic information

Entry
Database: EMDB / ID: EMD-29377
TitleAAV1 VP3 Only Capsid
Map data
Sample
  • Virus: Adeno-associated virus - 1
    • Protein or peptide: Capsid proteinCapsid
KeywordsAAV / gene therapy / capsid / VP3 / VIRUS LIKE PARTICLE
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 - 1
Methodsingle particle reconstruction / cryo EM / Resolution: 2.27 Å
AuthorsMietzsch M / McKenna R
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM082946 United States
CitationJournal: Mol Ther Methods Clin Dev / Year: 2023
Title: Production and characterization of an AAV1-VP3-only capsid: An analytical benchmark standard.
Authors: Mario Mietzsch / Weijing Liu / Ke Ma / Antonette Bennett / Austin R Nelson / Keely Gliwa / Paul Chipman / Xiaofeng Fu / Shane Bechler / Robert McKenna / Rosa Viner /
Abstract: Adeno-associated viruses (AAVs) are non-enveloped ssDNA icosahedral T = 1 viruses used as vectors for clinical gene delivery. Currently, there are over 200 AAV-related clinical trials and six ...Adeno-associated viruses (AAVs) are non-enveloped ssDNA icosahedral T = 1 viruses used as vectors for clinical gene delivery. Currently, there are over 200 AAV-related clinical trials and six approved biologics on the market. As such new analytical methods are continually being developed to characterize and monitor the quality and purity of manufactured AAV vectors, these include ion-exchange chromatography and Direct Mass Technology. However, these methods require homogeneous analytical standards with a high molecular weight standard comparable to the mass of an AAV capsid. Described here is the design, production, purification, characterization, and the cryo-electron microscopy structure of an AAV1-VP3-only capsid that fulfills this need as a calibrant to determine capsid mass, charge, homogeneity, and transgene packaging characteristics.
History
DepositionJan 5, 2023-
Header (metadata) releaseFeb 8, 2023-
Map releaseFeb 8, 2023-
UpdateJun 21, 2023-
Current statusJun 21, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_29377.png

Downloads & links

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Map

FileDownload / File: emd_29377.map.gz / Format: CCP4 / Size: 371.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.939 Å
Density
Contour LevelBy AUTHOR: 1.5
Minimum - Maximum-8.280195000000001 - 18.176079999999999
Average (Standard dev.)-0.000000000495646 (±0.99999994)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin-230-230-230
Dimensions460460460
Spacing460460460
CellA=B=C: 431.94 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_29377_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_29377_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Adeno-associated virus - 1

EntireName: Adeno-associated virus - 1
Components
  • Virus: Adeno-associated virus - 1
    • Protein or peptide: Capsid proteinCapsid

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Supramolecule #1: Adeno-associated virus - 1

SupramoleculeName: Adeno-associated virus - 1 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / NCBI-ID: 85106 / Sci species name: Adeno-associated virus - 1 / Virus type: VIRUS-LIKE PARTICLE / Virus isolate: SEROTYPE / Virus enveloped: No / Virus empty: Yes
Molecular weightTheoretical: 3.57 MDa
Virus shellShell ID: 1 / T number (triangulation number): 1

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Macromolecule #1: Capsid protein

MacromoleculeName: Capsid protein / type: protein_or_peptide / ID: 1 / Number of copies: 60 / Enantiomer: LEVO
Source (natural)Organism: Adeno-associated virus - 1
Molecular weightTheoretical: 58.299422 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: ADGVGNASGN WHCDSTWLGD RVITTSTRTW ALPTYNNHLY KQISSASTGA SNDNHYFGYS TPWGYFDFNR FHCHFSPRDW QRLINNNWG FRPKRLNFKL FNIQVKEVTT NDGVTTIANN LTSTVQVFSD SEYQLPYVLG SAHQGCLPPF PADVFMIPQY G YLTLNNGS ...String:
ADGVGNASGN WHCDSTWLGD RVITTSTRTW ALPTYNNHLY KQISSASTGA SNDNHYFGYS TPWGYFDFNR FHCHFSPRDW QRLINNNWG FRPKRLNFKL FNIQVKEVTT NDGVTTIANN LTSTVQVFSD SEYQLPYVLG SAHQGCLPPF PADVFMIPQY G YLTLNNGS QAVGRSSFYC LEYFPSQMLR TGNNFTFSYT FEEVPFHSSY AHSQSLDRLM NPLIDQYLYY LNRTQNQSGS AQ NKDLLFS RGSPAGMSVQ PKNWLPGPCY RQQRVSKTKT DNNNSNFTWT GASKYNLNGR ESIINPGTAM ASHKDDEDKF FPM SGVMIF GKESAGASNT ALDNVMITDE EEIKATNPVA TERFGTVAVN FQSSSTDPAT GDVHAMGALP GMVWQDRDVY LQGP IWAKI PHTDGHFHPS PLMGGFGLKN PPPQILIKNT PVPANPPAEF SATKFASFIT QYSTGQVSVE IEWELQKENS KRWNP EVQY TSNYAKSANV DFTVDNNGLY TEPRPIGTRY LTRPL

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 1.84 µm / Nominal defocus min: 0.48 µm
Image recordingFilm or detector model: DIRECT ELECTRON APOLLO (4k x 4k) / Average electron dose: 59.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 2.27 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 4418

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