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- PDB-7ltm: Hum8 capsid -

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Basic information

Entry
Database: PDB / ID: 7ltm
TitleHum8 capsid
ComponentsCapsid protein
KeywordsVIRUS / Icosahedral Capsid / AAV8 / Hum8 / capsid engineering / Adeno-associated virus / Parvovirus / Gene Therapy
Function / homology
Function and homology information


T=1 icosahedral viral capsid / structural molecule activity
Similarity search - Function
Phospholipase 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
Similarity search - Domain/homology
2'-DEOXYADENOSINE-5'-MONOPHOSPHATE / Capsid protein
Similarity search - Component
Biological speciesAdeno-associated virus - 8
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.49 Å
AuthorsMietzsch, M. / Agbandje-McKenna, M.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM082946 United States
CitationJournal: J Virol / Year: 2021
Title: Receptor Switching in Newly Evolved Adeno-associated Viruses.
Authors: L Patrick Havlik / Anshuman Das / Mario Mietzsch / Daniel K Oh / Jonathan Ark / Robert McKenna / Mavis Agbandje-McKenna / Aravind Asokan /
Abstract: Adeno-associated viruses utilize different glycans and the AAV receptor (AAVR) for cellular attachment and entry. Directed evolution has yielded new AAV variants; however, structure-function ...Adeno-associated viruses utilize different glycans and the AAV receptor (AAVR) for cellular attachment and entry. Directed evolution has yielded new AAV variants; however, structure-function correlates underlying their improved transduction are generally overlooked. Here, we report that infectious cycling of structurally diverse AAV surface loop libraries yields functionally distinct variants. Newly evolved variants show enhanced cellular binding, uptake, and transduction, but through distinct mechanisms. Using glycan-based and genome-wide CRISPR knockout screens, we discover that one AAV variant acquires the ability to recognize sulfated glycosaminoglycans, while another displays receptor switching from AAVR to integrin β1 (ITGB1). A previously evolved variant, AAVhum.8, preferentially utilizes the ITGB1 receptor over AAVR. Visualization of the AAVhum.8 capsid by cryoelectron microscopy at 2.49-Å resolution localizes the newly acquired integrin recognition motif adjacent to the AAVR footprint. These observations underscore the new finding that distinct AAV surface epitopes can be evolved to exploit different cellular receptors for enhanced transduction. Understanding how viruses interact with host cells through cell surface receptors is central to discovery and development of antiviral therapeutics, vaccines, and gene transfer vectors. Here, we demonstrate that distinct epitopes on the surface of adeno-associated viruses can be evolved by infectious cycling to recognize different cell surface carbohydrates and glycoprotein receptors and solve the three-dimensional structure of one such newly evolved AAV capsid, which provides a roadmap for designing viruses with improved attributes for gene therapy applications.
History
DepositionFeb 19, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 7, 2021Provider: repository / Type: Initial release
Revision 1.1Aug 18, 2021Group: Database references / Category: citation / citation_author / database_2
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.2Sep 22, 2021Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.title / _citation_author.identifier_ORCID
Revision 2.0May 29, 2024Group: Advisory / Atomic model ...Advisory / Atomic model / Data collection / Derived calculations / Non-polymer description
Category: atom_site / chem_comp ...atom_site / chem_comp / chem_comp_atom / chem_comp_bond / pdbx_entity_nonpoly / pdbx_nonpoly_scheme / pdbx_unobs_or_zero_occ_atoms / struct_site
Item: _atom_site.B_iso_or_equiv / _atom_site.Cartn_x ..._atom_site.B_iso_or_equiv / _atom_site.Cartn_x / _atom_site.Cartn_y / _atom_site.Cartn_z / _atom_site.auth_atom_id / _atom_site.auth_comp_id / _atom_site.label_atom_id / _atom_site.label_comp_id / _atom_site.type_symbol / _chem_comp.id / _chem_comp.mon_nstd_flag / _chem_comp.type / _pdbx_entity_nonpoly.comp_id / _pdbx_nonpoly_scheme.mon_id / _pdbx_nonpoly_scheme.pdb_mon_id / _pdbx_unobs_or_zero_occ_atoms.auth_atom_id / _pdbx_unobs_or_zero_occ_atoms.auth_comp_id / _pdbx_unobs_or_zero_occ_atoms.label_atom_id / _pdbx_unobs_or_zero_occ_atoms.label_comp_id / _struct_site.details / _struct_site.pdbx_auth_comp_id

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Structure visualization

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Structure viewerMolecule:
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Assembly

Deposited unit
A: Capsid protein
B: Capsid protein
C: Capsid protein
D: Capsid protein
E: Capsid protein
F: Capsid protein
G: Capsid protein
H: Capsid protein
I: Capsid protein
J: Capsid protein
K: Capsid protein
L: Capsid protein
M: Capsid protein
N: Capsid protein
O: Capsid protein
P: Capsid protein
Q: Capsid protein
R: Capsid protein
S: Capsid protein
T: Capsid protein
U: Capsid protein
V: Capsid protein
W: Capsid protein
X: Capsid protein
Y: Capsid protein
Z: Capsid protein
a: Capsid protein
b: Capsid protein
c: Capsid protein
d: Capsid protein
e: Capsid protein
f: Capsid protein
g: Capsid protein
h: Capsid protein
i: Capsid protein
j: Capsid protein
k: Capsid protein
l: Capsid protein
m: Capsid protein
n: Capsid protein
o: Capsid protein
p: Capsid protein
q: Capsid protein
r: Capsid protein
s: Capsid protein
t: Capsid protein
u: Capsid protein
v: Capsid protein
w: Capsid protein
x: Capsid protein
y: Capsid protein
z: Capsid protein
1: Capsid protein
2: Capsid protein
3: Capsid protein
4: Capsid protein
5: Capsid protein
6: Capsid protein
7: Capsid protein
8: Capsid protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)3,529,357120
Polymers3,509,48460
Non-polymers19,87360
Water00
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein ...
Capsid protein


Mass: 58491.398 Da / Num. of mol.: 60
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Adeno-associated virus - 8 / Production host: Homo sapiens (human) / References: UniProt: Q8JQF8
#2: Chemical...
ChemComp-D5M / 2'-DEOXYADENOSINE-5'-MONOPHOSPHATE


Mass: 331.222 Da / Num. of mol.: 60 / Source method: obtained synthetically / Formula: C10H14N5O6P
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Adeno-associated virus / Type: VIRUS / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Adeno-associated virus / Strain: Hum8
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293
Details of virusEmpty: NO / Enveloped: NO / Isolate: OTHER / Type: VIRION
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Cs: 2.7 mm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.10-2155_2155: / Classification: refinement
EM software
IDNameCategory
4cisTEMCTF correction
13cisTEM3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.49 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 275883 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.009256860
ELECTRON MICROSCOPYf_angle_d0.924350520
ELECTRON MICROSCOPYf_dihedral_angle_d8.814202800
ELECTRON MICROSCOPYf_chiral_restr0.0636420
ELECTRON MICROSCOPYf_plane_restr0.00646380

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