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- EMDB-21004: genome-containing AAV8 particles -

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Basic information

Entry
Database: EMDB / ID: EMD-21004
Titlegenome-containing AAV8 particles
Map data
SampleAdeno-associated virus != Adeno-associated virus - 8

Adeno-associated virus

  • Virus: Adeno-associated virus - 8
    • Protein or peptide: Capsid protein
  • DNA: DNA (5'-D(*CP*A)-3')
KeywordsAAV8 / capsid / genome / nucleotide binding pocket / VIRUS
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 - 8 / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.77 Å
AuthorsMietzsch M / Agbandje-McKenna M
CitationJournal: J Virol / Year: 2020
Title: Comparative Analysis of the Capsid Structures of AAVrh.10, AAVrh.39, and AAV8.
Authors: Mario Mietzsch / Candace Barnes / Joshua A Hull / Paul Chipman / Jun Xie / Nilakshee Bhattacharya / Duncan Sousa / Robert McKenna / Guangping Gao / Mavis Agbandje-McKenna /
Abstract: Adeno-associated viruses (AAVs) from clade E are often used as vectors in gene delivery applications. This clade includes rhesus isolate 10 (AAVrh.10) and 39 (AAVrh.39) which, unlike representative ...Adeno-associated viruses (AAVs) from clade E are often used as vectors in gene delivery applications. This clade includes rhesus isolate 10 (AAVrh.10) and 39 (AAVrh.39) which, unlike representative AAV8, are capable of crossing the blood-brain barrier (BBB), thereby enabling the delivery of therapeutic genes to the central nervous system. Here, the capsid structures of AAV8, AAVrh.10 and AAVrh.39 have been determined by cryo-electron microscopy and three-dimensional image reconstruction to 3.08-, 2.75-, and 3.39-Å resolution, respectively, to enable a direct structural comparison. AAVrh.10 and AAVrh.39 are 98% identical in amino acid sequence but only ∼93.5% identical to AAV8. However, the capsid structures of all three viruses are similar, with only minor differences observed in the previously described surface variable regions, suggesting that specific residues S269 and N472, absent in AAV8, may confer the ability to cross the BBB in AAVrh.10 and AAVrh.39. Head-to-head comparison of empty and genome-containing particles showed DNA ordered in the previously described nucleotide-binding pocket, supporting the suggested role of this pocket in DNA packaging for the The structural characterization of these viruses provides a platform for future vector engineering efforts toward improved gene delivery success with respect to specific tissue targeting, transduction efficiency, antigenicity, or receptor retargeting. Recombinant adeno-associated virus vectors (rAAVs), based on AAV8 and AAVrh.10, have been utilized in multiple clinical trials to treat different monogenetic diseases. The closely related AAVrh.39 has also shown promise As recently attained for other AAV biologics, e.g., Luxturna and Zolgensma, based on AAV2 and AAV9, respectively, the vectors in this study will likely gain U.S. Food and Drug Administration approval for commercialization in the near future. This study characterized the capsid structures of these clinical vectors at atomic resolution using cryo-electron microscopy and image reconstruction for comparative analysis. The analysis suggested two key residues, S269 and N472, as determinants of BBB crossing for AAVrh.10 and AAVrh.39, a feature utilized for central nervous system delivery of therapeutic genes. The structure information thus provides a platform for engineering to improve receptor retargeting or tissue specificity. These are important challenges in the field that need attention. Capsid structure information also provides knowledge potentially applicable for regulatory product approval.
History
DepositionNov 19, 2019-
Header (metadata) releaseDec 11, 2019-
Map releaseDec 11, 2019-
UpdateJun 12, 2024-
Current statusJun 12, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1.5
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 1.5
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6v10
  • Surface level: 1.5
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21004.png

Downloads & links

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Map

FileDownload / File: emd_21004.map.gz / Format: CCP4 / Size: 242.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)X (Row.)Y (Col.)
0.97 Å/pix.
x 399 pix.
= 387.03 Å		0.97 Å/pix.
x 399 pix.
= 387.03 Å		0.97 Å/pix.
x 399 pix.
= 387.03 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.97 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.0 / Movie #1: 1.5
Minimum - Maximum-8.260083 - 13.200024000000001
Average (Standard dev.)-0.000000002454535 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin-199-199-199
Dimensions399399399
Spacing399399399
CellA=B=C: 387.03 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.970.970.97
M x/y/z399399399
origin x/y/z0.0000.0000.000
length x/y/z387.030387.030387.030
α/β/γ90.00090.00090.000
start NX/NY/NZ-199-199-199
NX/NY/NZ399399399
MAP C/R/S213
start NC/NR/NS-199-199-199
NC/NR/NS399399399
D min/max/mean-8.26013.200-0.000

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Supplemental data

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Sample components

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Entire : Adeno-associated virus

EntireName: Adeno-associated virus
Components
  • Virus: Adeno-associated virus - 8
    • Protein or peptide: Capsid protein
  • DNA: DNA (5'-D(*CP*A)-3')

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Supramolecule #1: Adeno-associated virus - 8

SupramoleculeName: Adeno-associated virus - 8 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1 / NCBI-ID: 202813 / Sci species name: Adeno-associated virus - 8 / Sci species strain: AAV8 / Virus type: VIRION / Virus isolate: SEROTYPE / Virus enveloped: No / Virus empty: No

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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 - 8
Molecular weightTheoretical: 58.656512 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: GADGVGSSSG NWHCDSTWLG DRVITTSTRT WALPTYNNHL YKQISNGTSG GATNDNTYFG YSTPWGYFDF NRFHCHFSPR DWQRLINNN WGFRPKRLSF KLFNIQVKEV TQNEGTKTIA NNLTSTIQVF TDSEYQLPYV LGSAHQGCLP PFPADVFMIP Q YGYLTLNN ...String:
GADGVGSSSG NWHCDSTWLG DRVITTSTRT WALPTYNNHL YKQISNGTSG GATNDNTYFG YSTPWGYFDF NRFHCHFSPR DWQRLINNN WGFRPKRLSF KLFNIQVKEV TQNEGTKTIA NNLTSTIQVF TDSEYQLPYV LGSAHQGCLP PFPADVFMIP Q YGYLTLNN GSQAVGRSSF YCLEYFPSQM LRTGNNFQFT YTFEDVPFHS SYAHSQSLDR LMNPLIDQYL YYLSRTQTTG GT ANTQTLG FSQGGPNTMA NQAKNWLPGP CYRQQRVSTT TGQNNNSNFA WTAGTKYHLN GRNSLANPGI AMATHKDDEE RFF PSNGIL IFGKQNAARD NADYSDVMLT SEEEIKTTNP VATEEYGIVA DNLQQQNTAP QIGTVNSQGA LPGMVWQNRD VYLQ GPIWA KIPHTDGNFH PSPLMGGFGL KHPPPQILIK NTPVPADPPT TFNQSKLNSF ITQYSTGQVS VEIEWELQKE NSKRW NPEI QYTSNYYKST SVDFAVNTEG VYSEPRPIGT RYLTRNL

UniProtKB: Capsid protein

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Macromolecule #2: DNA (5'-D(*CP*A)-3')

MacromoleculeName: DNA (5'-D(*CP*A)-3') / type: dna / ID: 2 / Number of copies: 60 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 557.431 Da
SequenceString:
(DC)(DA)

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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
Image recordingFilm or detector model: DIRECT ELECTRON DE-20 (5k x 3k) / Average electron dose: 67.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 3.77 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 1072
Initial angle assignmentType: COMMON LINE
Final angle assignmentType: COMMON LINE

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