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

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Basic information

Entry
Database: EMDB / ID: 0535
TitleCryo-EM Reconstruction of Protease-Activateable Adeno-Associated Virus 9 (AAV9-L001)
Map dataProtease-Activateable Adeno-associated Virus 9 (AAV9-L001)
SampleAdeno-associated virus:
virus / Capsid protein VP1
Function / homologyParvovirus coat protein VP2 / Parvovirus coat protein VP1, N-terminal / Capsid/spike protein, ssDNA virus / Parvovirus coat protein VP1/VP2 / Parvovirus coat protein VP2 / Parvovirus coat protein VP1 / viral capsid / structural molecule activity / Capsid protein VP1
Function and homology information
SourceAdeno-associated virus
Methodsingle particle reconstruction / cryo EM / 3.16 Å resolution
AuthorsBennett AB / Agbandje-Mckenna M
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 / 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.
Validation ReportPDB-ID: 6nxe

SummaryFull reportAbout validation report
DateDeposition: Feb 8, 2019 / Header (metadata) release: Mar 6, 2019 / Map release: Mar 13, 2019 / Last update: Mar 20, 2019

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Structure visualization

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

Downloads & links

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Map

Fileemd_0535.map.gz (map file in CCP4 format, 277707 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)X (Row.)Y (Col.)
411 pix
0.97 Å/pix.
= 400.314 Å
411 pix
0.97 Å/pix.
= 400.314 Å
411 pix
0.97 Å/pix.
= 400.314 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.974 Å
Density
Contour Level:1.0 (by author), 1 (movie #1):
Minimum - Maximum-6.825892 - 17.40653
Average (Standard dev.)0.000000003102391 (1.0)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderYXZ
Dimensions411411411
Origin-205.0-205.0-205.0
Limit205.0205.0205.0
Spacing411411411
CellA=B=C: 400.314 Å
α=β=γ: 90.0 deg.

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 / Number of components: 2

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Component #1: virus, Adeno-associated virus

VirusName: Adeno-associated virus / Class: VIRION / Empty: No / Enveloped: No / Isolate: SEROTYPE
SpeciesSpecies: Adeno-associated virus / Strain: 9-L001
Source (engineered)Expression System: Homo sapiens (human)

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Component #2: protein, Capsid protein VP1

ProteinName: Capsid protein VP1 / Number of Copies: 60 / Recombinant expression: No
MassTheoretical: 60.939168 kDa
SourceSpecies: Adeno-associated virus
Source (engineered)Expression System: Homo sapiens (human)

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.5 mg/ml / pH: 7.4
Support filmunspecified
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 2 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: DIRECT ELECTRON DE-20 (5k x 3k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 114044
3D reconstructionResolution: 3.16 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement space: REAL
Input PDB model: 3UX1
Chain ID: A
Output model

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