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- PDB-8jif: Cryo-EM Structure of 3-axis block of AAV9P31-Car4 complex -

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Basic information

Entry
Database: PDB / ID: 8jif
TitleCryo-EM Structure of 3-axis block of AAV9P31-Car4 complex
Components
  • Capsid protein VP1
  • Carbonic anhydrase 4
KeywordsVIRAL PROTEIN/LYASE / receptor / aav9p31 / Carbonic anhydrases iv / Block-based reconstruction / VIRUS / VIRAL PROTEIN-LYASE complex
Function / homology
Function and homology information


Erythrocytes take up carbon dioxide and release oxygen / Erythrocytes take up oxygen and release carbon dioxide / Reversible hydration of carbon dioxide / regulation of pH / bicarbonate transport / T=1 icosahedral viral capsid / transport vesicle membrane / endoplasmic reticulum-Golgi intermediate compartment / rough endoplasmic reticulum / secretory granule membrane ...Erythrocytes take up carbon dioxide and release oxygen / Erythrocytes take up oxygen and release carbon dioxide / Reversible hydration of carbon dioxide / regulation of pH / bicarbonate transport / T=1 icosahedral viral capsid / transport vesicle membrane / endoplasmic reticulum-Golgi intermediate compartment / rough endoplasmic reticulum / secretory granule membrane / sarcoplasmic reticulum / carbonic anhydrase / brush border membrane / carbonate dehydratase activity / carbon dioxide transport / trans-Golgi network / sarcolemma / one-carbon metabolic process / apical plasma membrane / external side of plasma membrane / structural molecule activity / perinuclear region of cytoplasm / zinc ion binding / extracellular exosome / membrane / plasma membrane
Similarity search - Function
Carbonic anhydrase, CA4/CA15 / 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 / Carbonic anhydrase, alpha-class, conserved site / Alpha-carbonic anhydrases signature. / Carbonic anhydrase, alpha-class ...Carbonic anhydrase, CA4/CA15 / 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 / Carbonic anhydrase, alpha-class, conserved site / Alpha-carbonic anhydrases signature. / Carbonic anhydrase, alpha-class / Alpha carbonic anhydrase domain / Alpha carbonic anhydrase domain superfamily / Eukaryotic-type carbonic anhydrase / Alpha-carbonic anhydrases profile. / Eukaryotic-type carbonic anhydrase
Similarity search - Domain/homology
Carbonic anhydrase 4 / Capsid protein VP1
Similarity search - Component
Biological speciesMus musculus (house mouse)
Adeno-associated virus 9
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.28 Å
AuthorsZhang, R. / Liu, Y. / Lou, Z.
Funding support China, 3items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31971126 China
National Natural Science Foundation of China (NSFC)U20A20135 China
National Natural Science Foundation of China (NSFC)32301011 China
CitationJournal: PLoS Pathog / Year: 2024
Title: Structural basis of the recognition of adeno-associated virus by the neurological system-related receptor carbonic anhydrase IV.
Authors: Ran Zhang / Yixiao Liu / Fengxi Yu / Guangxue Xu / Lili Li / Baobin Li / Zhiyong Lou /
Abstract: Carbonic anhydrase IV (Car4) is a newly identified receptor that allows adeno-associated virus (AAV) 9P31 to cross the blood-brain barrier and achieve efficient infection in the central nervous ...Carbonic anhydrase IV (Car4) is a newly identified receptor that allows adeno-associated virus (AAV) 9P31 to cross the blood-brain barrier and achieve efficient infection in the central nervous system (CNS) in mouse models. However, the molecular mechanism by which engineered AAV capsids with 7-mer insertion in the variable region (VR) VIII recognize these novel cellular receptors is unknown. Here we report the cryo-EM structures of AAV9P31 and its complex with Mus musculus Car4 at atomic resolution by utilizing the block-based reconstruction (BBR) method. The structures demonstrated that Car4 binds to the protrusions at 3-fold axes of the capsid. The inserted 7-mer extends into a hydrophobic region near the catalytic center of Car4 to form stable interactions. Mutagenesis studies also identified the key residues in Car4 responsible for the AAV9P31 interaction. These findings provide new insights into the novel receptor recognition mechanism of AAV generated by directed evolution and highlight the application of the BBR method to studying the virus-receptor molecular mechanism.
History
DepositionMay 26, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 31, 2024Provider: repository / Type: Initial release
Revision 1.1Feb 28, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
R: Carbonic anhydrase 4
A: Capsid protein VP1
B: Capsid protein VP1
C: Capsid protein VP1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)207,2335
Polymers207,1674
Non-polymers651
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Carbonic anhydrase 4 / Carbonate dehydratase IV / Carbonic anhydrase IV / CA-IV


Mass: 29281.367 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Ca4, Car4
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: Q64444, carbonic anhydrase
#2: Protein Capsid protein VP1


Mass: 59295.277 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Adeno-associated virus 9 / Gene: cap / Production host: Homo sapiens (human) / References: UniProt: Q6JC40
#3: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Zn / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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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: 3-fold axes block of AAV9P31 and Car4 complex aat 2.28 angstrom
Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
11Mus musculus (house mouse)10090
21Adeno-associated virus 9235455
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
11Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)866768
21Homo sapiens (human)9606
Buffer solutionpH: 8 / Details: 20mM Tris-Cl, pH8.0; 150mM NaCl.
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMTris-ClTris-Cl1
2150 mMNaClNaCl1
SpecimenConc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 280 K

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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 / Nominal defocus max: 1800 nm / Nominal defocus min: 600 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K2 QUANTUM (4k x 4k)

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Processing

EM software
IDNameCategory
2SerialEMimage acquisition
4CTFFINDCTF correction
10RELIONinitial Euler assignment
11RELIONfinal Euler assignment
CTF correctionType: NONE
3D reconstructionResolution: 2.28 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 13604676 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0022107
ELECTRON MICROSCOPYf_angle_d0.512852
ELECTRON MICROSCOPYf_dihedral_angle_d11.074799
ELECTRON MICROSCOPYf_chiral_restr0.043303
ELECTRON MICROSCOPYf_plane_restr0.004370

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