+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-20624 | |||||||||
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Title | Empty AAV3B | |||||||||
Map data | Empty AAV3B (No genome) | |||||||||
Sample | Parvoviridae != Adeno-associated virus 3B Parvoviridae
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Biological species | Homo sapiens (human) / Adeno-associated virus 3B | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.26 Å | |||||||||
Authors | Subramanian S / Hafenstein S | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Hum Gene Ther / Year: 2019 Title: Filling Adeno-Associated Virus Capsids: Estimating Success by Cryo-Electron Microscopy. Authors: Suriyasri Subramanian / Anna C Maurer / Carol M Bator / Alexander M Makhov / James F Conway / Kevin B Turner / James H Marden / Luk H Vandenberghe / Susan L Hafenstein / Abstract: Adeno-associated viruses (AAVs) have been employed successfully as gene therapy vectors in treating various genetic diseases for almost two decades. However, transgene packaging is usually imperfect, ...Adeno-associated viruses (AAVs) have been employed successfully as gene therapy vectors in treating various genetic diseases for almost two decades. However, transgene packaging is usually imperfect, and developing a rapid and accurate method for measuring the proportion of DNA encapsidation is an important step for improving the downstream process of large scale vector production. In this study, we used two-dimensional class averages and three-dimensional classes, intermediate outputs in the single particle cryo-electron microscopy (cryo-EM) image reconstruction pipeline, to determine the proportion of DNA-packaged and empty capsid populations. Two different preparations of AAV3 were analyzed to estimate the minimum number of particles required to be sampled by cryo-EM in order for robust calculation of the proportion of the full versus empty capsids in any given sample. Cost analysis applied to the minimum amount of data required for a valid ratio suggests that cryo-EM is an effective approach to analyze vector preparations. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_20624.map.gz | 181 MB | EMDB map data format | |
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Header (meta data) | emd-20624-v30.xml emd-20624.xml | 12.5 KB 12.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_20624_fsc.xml | 13.1 KB | Display | FSC data file |
Images | emd_20624.png | 232.7 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-20624 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-20624 | HTTPS FTP |
-Validation report
Summary document | emd_20624_validation.pdf.gz | 77.8 KB | Display | EMDB validaton report |
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Full document | emd_20624_full_validation.pdf.gz | 76.9 KB | Display | |
Data in XML | emd_20624_validation.xml.gz | 495 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-20624 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-20624 | HTTPS FTP |
-Related structure data
Related structure data | C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_20624.map.gz / Format: CCP4 / Size: 193.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Empty AAV3B (No genome) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.136 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Parvoviridae
Entire | Name: Parvoviridae (virus) |
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Components |
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-Supramolecule #1: Adeno-associated virus 3B
Supramolecule | Name: Adeno-associated virus 3B / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / NCBI-ID: 68742 / Sci species name: Adeno-associated virus 3B / Virus type: VIRUS-LIKE PARTICLE / Virus isolate: OTHER / Virus enveloped: No / Virus empty: Yes |
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Host (natural) | Organism: Mus spretus (western wild mouse) |
Host system | Organism: Homo sapiens (human) / Recombinant cell: HEK |
Virus shell | Shell ID: 1 / Diameter: 250.0 Å / T number (triangulation number): 1 |
-Macromolecule #1: AAV3B
Macromolecule | Name: AAV3B / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MAADGYLPDW LEDNLSEGIR EWWALKPGVP QPKANQQHQD NRRGLVLPGY KYLGPGNGLD KGEPVNEAD AAALEHDKAY DQQLKAGDNP YLKYNHADAE FQERLQEDTS FGGNLGRAVF Q AKKRILEP LGLVEEAAKT APGKKRPVDQ SPQEPDSSSG VGKSGKQPAR ...String: MAADGYLPDW LEDNLSEGIR EWWALKPGVP QPKANQQHQD NRRGLVLPGY KYLGPGNGLD KGEPVNEAD AAALEHDKAY DQQLKAGDNP YLKYNHADAE FQERLQEDTS FGGNLGRAVF Q AKKRILEP LGLVEEAAKT APGKKRPVDQ SPQEPDSSSG VGKSGKQPAR KRLNFGQTGD SE SVPDPQP LGEPPAAPTS LGSNTMASGG GAPMADNNEG ADGVGNSSGN WHCDSQWLGD RVI TTSTRT WALPTYNNHL YKQISSQSGA SNDNHYFGYS TPWGYFDFNR FHCHFSPRDW QRLI NNNWG FRPKKLSFKL FNIQVKEVTQ NDGTTTIANN LTSTVQVFTD SEYQLPYVLG SAHQG CLPP FPADVFMVPQ YGYLTLNNGS QAVGRSSFYC LEYFPSQMLR TGNNFQFSYT FEDVPF HSS YAHSQSLDRL MNPLIDQYLY YLNRTQGTTS GTTNQSRLLF SQAGPQSMSL QARNWLP GP CYRQQRLSKT ANDNNNSNFP WTAASKYHLN GRDSLVNPGP AMASHKDDEE KFFPMHGN L IFGKEGTTAS NAELDNVMIT DEEEIRTTNP VATEQYGTVA NNLQSSNTAP TTRTVNDQG ALPGMVWQDR DVYLQGPIWA KIPHTDGHFH PSPLMGGFGL KHPPPQIMIK NTPVPANPPT TFSPAKFAS FITQYSTGQV SVEIEWELQK ENSKRWNPEI QYTSNYNKSV NVDFTVDTNG V YSEPRPIG TRYLTRNL |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.4 Component:
Details: Phosphate buffered saline pH 7.4 | |||||||||||||||
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Grid | Details: unspecified | |||||||||||||||
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number real images: 2082 / Average electron dose: 45.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: DARK FIELD |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |