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- EMDB-0535: Cryo-EM Reconstruction of Protease-Activateable Adeno-Associated ... -

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Basic information

Entry
Database: EMDB / ID: EMD-0535
TitleCryo-EM Reconstruction of Protease-Activateable Adeno-Associated Virus 9 (AAV9-L001)
Map dataProtease-Activateable Adeno-associated Virus 9 (AAV9-L001)
Sample
  • Virus: Adeno-associated virus
    • Protein or peptide: Capsid protein VP1
KeywordsAAV / activatable / gene therapy / provector / 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: 3.16 Å
AuthorsBennett AB / Agbandje-Mckenna M
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI)R21HL126053, R01HL138126, and R01CA207497 United States
CitationJournal: Mol Ther / Year: 2019
Title: Protease-Activatable Adeno-Associated Virus Vector for Gene Delivery to Damaged Heart Tissue.
Authors: Caitlin M Guenther / Mitchell J Brun / Antonette D Bennett / Michelle L Ho / Weitong Chen / Banghe Zhu / Michael Lam / Momona Yamagami / Sunkuk Kwon / Nilakshee Bhattacharya / Duncan Sousa / ...Authors: Caitlin M Guenther / Mitchell J Brun / Antonette D Bennett / Michelle L Ho / Weitong Chen / Banghe Zhu / Michael Lam / Momona Yamagami / Sunkuk Kwon / Nilakshee Bhattacharya / Duncan Sousa / Annicka C Evans / Julie Voss / Eva M Sevick-Muraca / Mavis Agbandje-McKenna / Junghae Suh /
Abstract: Adeno-associated virus (AAV) has emerged as a promising gene delivery vector because of its non-pathogenicity, simple structure and genome, and low immunogenicity compared to other viruses. However, ...Adeno-associated virus (AAV) has emerged as a promising gene delivery vector because of its non-pathogenicity, simple structure and genome, and low immunogenicity compared to other viruses. However, its adoption as a safe and effective delivery vector for certain diseases relies on altering its tropism to deliver transgenes to desired cell populations. To this end, we have developed a protease-activatable AAV vector, named provector, that responds to elevated extracellular protease activity commonly found in diseased tissue microenvironments. The AAV9-based provector is initially inactive, but then it can be switched on by matrix metalloproteinases (MMP)-2 and -9. Cryo-electron microscopy and image reconstruction reveal that the provector capsid is structurally similar to that of AAV9, with a flexible peptide insertion at the top of the 3-fold protrusions. In an in vivo model of myocardial infarction (MI), the provector is able to deliver transgenes site specifically to high-MMP-activity regions of the damaged heart, with concomitant decreased delivery to many off-target organs, including the liver. The AAV provector may be useful in the future for enhanced delivery of transgenes to sites of cardiac damage.
History
DepositionFeb 8, 2019-
Header (metadata) releaseMar 6, 2019-
Map releaseMar 13, 2019-
UpdateMar 20, 2024-
Current statusMar 20, 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-6nxe
  • Surface level: 1
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6nxe
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0535.png

Downloads & links

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Map

FileDownload / File: emd_0535.map.gz / Format: CCP4 / Size: 264.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationProtease-Activateable Adeno-associated Virus 9 (AAV9-L001)
Voxel sizeX=Y=Z: 0.974 Å
Density
Contour LevelBy AUTHOR: 1.0 / Movie #1: 1
Minimum - Maximum-6.825892 - 17.40653
Average (Standard dev.)0.000000003102391 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin-205-205-205
Dimensions411411411
Spacing411411411
CellA=B=C: 400.314 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.9740.9740.974
M x/y/z411411411
origin x/y/z0.0000.0000.000
length x/y/z400.314400.314400.314
α/β/γ90.00090.00090.000
start NX/NY/NZ-205-205-205
NX/NY/NZ411411411
MAP C/R/S213
start NC/NR/NS-205-205-205
NC/NR/NS411411411
D min/max/mean-6.82617.4070.000

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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

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Supramolecule #1: Adeno-associated virus

SupramoleculeName: Adeno-associated virus / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / NCBI-ID: 272636 / Sci species name: Adeno-associated virus / Sci species strain: 9-L001 / Virus type: VIRION / Virus isolate: SEROTYPE / Virus enveloped: No / Virus empty: No
Virus shellShell ID: 1 / Diameter: 260.0 Å / T number (triangulation number): 1

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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: 60.939168 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DGVGSSSGNW HCDSQWLGDR VITTSTRTWA LPTYNNHLYK QISNSTSGGS SNDNAYFGYS TPWGYFDFNR FHCHFSPRDW QRLINNNWG FRPKRLNFKL FNIQVKEVTD NNGVKTIANN LTSTVQVFTD SDYQLPYVLG SAHEGCLPPF PADVFMIPQY G YLTLNDGS ...String:
DGVGSSSGNW HCDSQWLGDR VITTSTRTWA LPTYNNHLYK QISNSTSGGS SNDNAYFGYS TPWGYFDFNR FHCHFSPRDW QRLINNNWG FRPKRLNFKL FNIQVKEVTD NNGVKTIANN LTSTVQVFTD SDYQLPYVLG SAHEGCLPPF PADVFMIPQY G YLTLNDGS QAVGRSSFYC LEYFPSQMLR TGNNFQFSYE FENVPFHSSY AHSQSLDRLM NPLIDQYLYY LSKTINGAGV PM SMRGGGD DDDGVPMSMR GGGASGQNQQ TLKFSVAGPS NMAVQGRNYI PGPSYRQQRV STTVTQNNNS EFAWPGASSW ALN GRNSLM NPGPAMASHK EGEDRFFPLS GSLIFGKQGT GRDNVDADKV MITNEEEIKT TNPVATESYG QVATNHQSAQ AQAQ TGWVQ NQGILPGMVW QDRDVYLQGP IWAKIPHTDG NFHPSPLMGG FGMKHPPPQI LIKNTPVPAD PPTAFNKDKL NSFIT QYST GQVSVEIEWE LQKENSKRWN PEIQYTSNYY KSNNVEFAVN TEGVYSEPRP IGTRYLTRNL

UniProtKB: Capsid protein VP1

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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

Concentration0.5 mg/mL
BufferpH: 7.4 / Component - Formula: PBS-MK / Component - Name: TD Buffer
GridDetails: unspecified
VitrificationCryogen name: ETHANE
DetailsThe sample was monodisperse.

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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
Image recordingFilm or detector model: DIRECT ELECTRON DE-20 (5k x 3k) / Average electron dose: 20.0 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: PROJECTION MATCHING
Projection matching processing - Number reference projections: 100
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.16 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 114044

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Atomic model buiding 1

Initial modelPDB ID:

3ux1


Chain - Chain ID: A / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL
Output model

PDB-6nxe:
Cryo-EM Reconstruction of Protease-Activateable Adeno-Associated Virus 9 (AAV9-L001)

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