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- EMDB-21011: Genome-containing AAVrh.10 -

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Basic information

Entry
Database: EMDB / ID: EMD-21011
TitleGenome-containing AAVrh.10
Map dataGenome-containing AAVrh.10
Sample
  • Virus: Adeno-associated virus
    • Protein or peptide: Capsid protein VP1
  • Ligand: 2'-DEOXYCYTIDINE-5'-MONOPHOSPHATE
  • Ligand: 2'-DEOXYADENOSINE-5'-MONOPHOSPHATE
KeywordsAAVrh.10 / 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 VP1
Function and homology information
Biological speciesAdeno-associated virus
Methodsingle particle reconstruction / cryo EM / Resolution: 2.98 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 20, 2019-
Header (metadata) releaseDec 11, 2019-
Map releaseDec 11, 2019-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1
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  • Surface view colored by radius
  • Surface level: 1
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  • Surface view with fitted model
  • Atomic models: PDB-6v1g
  • Surface level: 1
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6v1g
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21011.png

Downloads & links

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Map

FileDownload / File: emd_21011.map.gz / Format: CCP4 / Size: 246 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationGenome-containing AAVrh.10
Voxel sizeX=Y=Z: 0.975 Å
Density
Contour LevelBy AUTHOR: 1.0 / Movie #1: 1
Minimum - Maximum-9.471681999999999 - 19.450279999999999
Average (Standard dev.)-0.000000001299121 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin-200-200-200
Dimensions401401401
Spacing401401401
CellA=B=C: 390.975 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.9750.9750.975
M x/y/z401401401
origin x/y/z0.0000.0000.000
length x/y/z390.975390.975390.975
α/β/γ90.00090.00090.000
start NX/NY/NZ-200-200-200
NX/NY/NZ401401401
MAP C/R/S213
start NC/NR/NS-200-200-200
NC/NR/NS401401401
D min/max/mean-9.47219.450-0.000

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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
    • Protein or peptide: Capsid protein VP1
  • Ligand: 2'-DEOXYCYTIDINE-5'-MONOPHOSPHATE
  • Ligand: 2'-DEOXYADENOSINE-5'-MONOPHOSPHATE

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Supramolecule #1: Adeno-associated virus

SupramoleculeName: Adeno-associated virus / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1 / NCBI-ID: 272636 / Sci species name: Adeno-associated virus / Virus type: VIRION / Virus isolate: SEROTYPE / Virus enveloped: No / Virus empty: No

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Macromolecule #1: Capsid protein VP1

MacromoleculeName: Capsid protein VP1 / type: protein_or_peptide / ID: 1 / Number of copies: 60 / Enantiomer: LEVO
Source (natural)Organism: Adeno-associated virus
Molecular weightTheoretical: 58.42841 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: ADGVGSSSGN WHCDSTWLGD RVITTSTRTW ALPTYNNHLY KQISNGTSGG STNDNTYFGY STPWGYFDFN RFHCHFSPRD WQRLINNNW GFRPKRLNFK LFNIQVKEVT QNEGTKTIAN NLTSTIQVFT DSEYQLPYVL GSAHQGCLPP FPADVFMIPQ Y GYLTLNNG ...String:
ADGVGSSSGN WHCDSTWLGD RVITTSTRTW ALPTYNNHLY KQISNGTSGG STNDNTYFGY STPWGYFDFN RFHCHFSPRD WQRLINNNW GFRPKRLNFK LFNIQVKEVT QNEGTKTIAN NLTSTIQVFT DSEYQLPYVL GSAHQGCLPP FPADVFMIPQ Y GYLTLNNG SQAVGRSSFY CLEYFPSQML RTGNNFEFSY QFEDVPFHSS YAHSQSLDRL MNPLIDQYLY YLSRTQSTGG TA GTQQLLF SQAGPNNMSA QAKNWLPGPC YRQQRVSTTL SQNNNSNFAW TGATKYHLNG RDSLVNPGVA MATHKDDEER FFP SSGVLM FGKQGAGKDN VDYSSVMLTS EEEIKTTNPV ATEQYGVVAD NLQQQNAAPI VGAVNSQGAL PGMVWQNRDV YLQG PIWAK IPHTDGNFHP SPLMGGFGLK HPPPQILIKN TPVPADPPTT FSQAKLASFI TQYSTGQVSV EIEWELQKEN SKRWN PEIQ YTSNYYKSTN VDFAVNTDGT YSEPRPIGTR YLTRNL

UniProtKB: Capsid protein VP1

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Macromolecule #2: 2'-DEOXYCYTIDINE-5'-MONOPHOSPHATE

MacromoleculeName: 2'-DEOXYCYTIDINE-5'-MONOPHOSPHATE / type: ligand / ID: 2 / Number of copies: 60 / Formula: DC
Molecular weightTheoretical: 307.197 Da
Chemical component information

ChemComp-DC:
2'-DEOXYCYTIDINE-5'-MONOPHOSPHATE / dCMP*YM / Deoxycytidine monophosphate

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Macromolecule #3: 2'-DEOXYADENOSINE-5'-MONOPHOSPHATE

MacromoleculeName: 2'-DEOXYADENOSINE-5'-MONOPHOSPHATE / type: ligand / ID: 3 / Number of copies: 60 / Formula: DA
Molecular weightTheoretical: 331.222 Da
Chemical component information

ChemComp-D5M:
2'-DEOXYADENOSINE-5'-MONOPHOSPHATE / Deoxyadenosine monophosphate

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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
GridDetails: unspecified
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
Image recordingFilm or detector model: DIRECT ELECTRON DE-64 (8k x 8k) / Average electron dose: 60.0 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: COMMON LINE
Final angle assignmentType: COMMON LINE
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 2.98 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 82463

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