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- EMDB-34412: Cryo-EM structure of APOBEC3G-Vif complex -

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Basic information

Entry
Database: EMDB / ID: EMD-34412
TitleCryo-EM structure of APOBEC3G-Vif complex
Map data
Sample
  • Complex: A3G-VC-RNA complex
    • Complex: APOVEC3G
      • Protein or peptide: APOBEC3G
      • Protein or peptide: Core binding factor beta
    • Complex: Vif
      • Protein or peptide: Viral infectivity factor
    • Complex: RNA
      • RNA: RNA (5'-R(*CP*GP*GP*UP*UP*GP*AP*UP*CP*GP*UP*UP*UP*UP*AP*AP*CP*AP*A)-3')
  • Ligand: ZINC ION
  • Ligand: CHLORIDE ION
KeywordsHuman antiviral protein HIV / ANTIVIRAL PROTEIN
Function / homology
Function and homology information


RUNX3 regulates RUNX1-mediated transcription / RUNX1 regulates transcription of genes involved in BCR signaling / RUNX1 regulates transcription of genes involved in interleukin signaling / RUNX2 regulates bone development / core-binding factor complex / RUNX1 regulates expression of components of tight junctions / positive regulation of CD8-positive, alpha-beta T cell differentiation / RUNX2 regulates chondrocyte maturation / negative regulation of CD4-positive, alpha-beta T cell differentiation / lymphocyte differentiation ...RUNX3 regulates RUNX1-mediated transcription / RUNX1 regulates transcription of genes involved in BCR signaling / RUNX1 regulates transcription of genes involved in interleukin signaling / RUNX2 regulates bone development / core-binding factor complex / RUNX1 regulates expression of components of tight junctions / positive regulation of CD8-positive, alpha-beta T cell differentiation / RUNX2 regulates chondrocyte maturation / negative regulation of CD4-positive, alpha-beta T cell differentiation / lymphocyte differentiation / RUNX1 and FOXP3 control the development of regulatory T lymphocytes (Tregs) / RUNX2 regulates genes involved in cell migration / RUNX2 regulates genes involved in differentiation of myeloid cells / Transcriptional regulation by RUNX2 / RUNX1 regulates transcription of genes involved in differentiation of keratinocytes / myeloid cell differentiation / RUNX3 Regulates Immune Response and Cell Migration / definitive hemopoiesis / RUNX1 regulates transcription of genes involved in differentiation of myeloid cells / Regulation of RUNX1 Expression and Activity / RUNX1 regulates transcription of genes involved in WNT signaling / RUNX1 regulates estrogen receptor mediated transcription / RUNX1 interacts with co-factors whose precise effect on RUNX1 targets is not known / RUNX2 regulates osteoblast differentiation / RUNX3 regulates p14-ARF / cell maturation / Regulation of RUNX3 expression and activity / RUNX1 regulates genes involved in megakaryocyte differentiation and platelet function / Transcriptional regulation of granulopoiesis / protein polyubiquitination / osteoblast differentiation / Regulation of RUNX2 expression and activity / RUNX1 regulates transcription of genes involved in differentiation of HSCs / Estrogen-dependent gene expression / sequence-specific DNA binding / transcription by RNA polymerase II / transcription coactivator activity / regulation of transcription by RNA polymerase II / negative regulation of transcription by RNA polymerase II / positive regulation of transcription by RNA polymerase II / nucleoplasm / membrane
Similarity search - Function
Core-binding factor, beta subunit / Core-binding factor, beta subunit superfamily / Core binding factor beta subunit
Similarity search - Domain/homology
Core-binding factor subunit beta
Similarity search - Component
Biological speciesHuman Immunodeficiency virus 1 / Homo sapiens (human) / Human immunodeficiency virus 1 / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsKouno T / Shibata S / Hyun J / Kim TG / Wolf M
Funding support Japan, 1 items
OrganizationGrant numberCountry
Japan Agency for Medical Research and Development (AMED)JP18am0101076 Japan
CitationJournal: Nat Commun / Year: 2023
Title: Structural insights into RNA bridging between HIV-1 Vif and antiviral factor APOBEC3G.
Authors: Takahide Kouno / Satoshi Shibata / Megumi Shigematsu / Jaekyung Hyun / Tae Gyun Kim / Hiroshi Matsuo / Matthias Wolf /
Abstract: Great effort has been devoted to discovering the basis of A3G-Vif interaction, the key event of HIV's counteraction mechanism to evade antiviral innate immune response. Here we show reconstitution of ...Great effort has been devoted to discovering the basis of A3G-Vif interaction, the key event of HIV's counteraction mechanism to evade antiviral innate immune response. Here we show reconstitution of the A3G-Vif complex and subsequent A3G ubiquitination in vitro and report the cryo-EM structure of the A3G-Vif complex at 2.8 Å resolution using solubility-enhanced variants of A3G and Vif. We present an atomic model of the A3G-Vif interface, which assembles via known amino acid determinants. This assembly is not achieved by protein-protein interaction alone, but also involves RNA. The cryo-EM structure and in vitro ubiquitination assays identify an adenine/guanine base preference for the interaction and a unique Vif-ribose contact. This establishes the biological significance of an RNA ligand. Further assessment of interactions between A3G, Vif, and RNA ligands show that the A3G-Vif assembly and subsequent ubiquitination can be controlled by amino acid mutations at the interface or by polynucleotide modification, suggesting that a specific chemical moiety would be a promising pharmacophore to inhibit the A3G-Vif interaction.
History
DepositionSep 29, 2022-
Header (metadata) releaseJul 19, 2023-
Map releaseJul 19, 2023-
UpdateJul 3, 2024-
Current statusJul 3, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_34412.png

Downloads & links

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Map

FileDownload / File: emd_34412.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 256 pix.
= 215.04 Å		0.84 Å/pix.
x 256 pix.
= 215.04 Å		0.84 Å/pix.
x 256 pix.
= 215.04 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.84 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 4.0
Minimum - Maximum-7.1680865 - 20.789470000000001
Average (Standard dev.)0.00000000052769 (±0.9999998)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 215.04 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_34412_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_34412_half_map_1.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Half map: #2

Fileemd_34412_half_map_2.map
Projections & Slices
AxesZYX

Projections

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Histogram

Histogram (log scale)

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Sample components

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Entire : A3G-VC-RNA complex

EntireName: A3G-VC-RNA complex
Components
  • Complex: A3G-VC-RNA complex
    • Complex: APOVEC3G
      • Protein or peptide: APOBEC3G
      • Protein or peptide: Core binding factor beta
    • Complex: Vif
      • Protein or peptide: Viral infectivity factor
    • Complex: RNA
      • RNA: RNA (5'-R(*CP*GP*GP*UP*UP*GP*AP*UP*CP*GP*UP*UP*UP*UP*AP*AP*CP*AP*A)-3')
  • Ligand: ZINC ION
  • Ligand: CHLORIDE ION

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Supramolecule #1: A3G-VC-RNA complex

SupramoleculeName: A3G-VC-RNA complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4

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Supramolecule #2: APOVEC3G

SupramoleculeName: APOVEC3G / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1, #3
Source (natural)Organism: Human Immunodeficiency virus 1

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Supramolecule #3: Vif

SupramoleculeName: Vif / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #4: RNA

SupramoleculeName: RNA / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #4

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Macromolecule #1: APOBEC3G

MacromoleculeName: APOBEC3G / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 43.857266 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: GPGHMDPDTF SYNFNNRPIL SRRNTVWLCY EVERLDNGTW VKMDQHRGQV YSELKYHPEM RFLSLVSKWK LHRDQEYEVT WYISWSPCT KCARDMATFL QENTHVTLTI FVARLYYFWD PDYQEALRSL AQAGATIKIM NYDEFQHCWS KFVYSQGAPF Q PWDGLDEY ...String:
GPGHMDPDTF SYNFNNRPIL SRRNTVWLCY EVERLDNGTW VKMDQHRGQV YSELKYHPEM RFLSLVSKWK LHRDQEYEVT WYISWSPCT KCARDMATFL QENTHVTLTI FVARLYYFWD PDYQEALRSL AQAGATIKIM NYDEFQHCWS KFVYSQGAPF Q PWDGLDEY SQALSGMLGE ILRHSMDPPT FTFNFNNEPW VRGRHETYLC YEVERMHNDT WVKLNQRRGF LANQAPHKHG FL EGRHAEL CFLDVIPFWK LDLDQDYRVT CFTSWSPCFS CAQEMAKFIS KNKHVSLCIK TARIYDDQGR AQEGLRTLAE AGA KISIMT YSEFKHCWDT FVDHQGAPFQ PWDGLDEHSQ DLSGRLRAIL QNQEN

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Macromolecule #2: Viral infectivity factor

MacromoleculeName: Viral infectivity factor / type: protein_or_peptide / ID: 2 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightTheoretical: 18.354801 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MGSSHHHHHH SQDPMENRWQ VMIVWQVDRM RINTWKRLVK HHMYISRKAK DWFYRHHYES TNPKISSEVH IPLGDAKLVI TTYWGLHTG ERDWHLGQGV SIEWRKKRYS TQVDPDLADQ LIHLHYFDEA SEGSQIKPPL PSVRKLTEDR WNK

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Macromolecule #3: Core binding factor beta

MacromoleculeName: Core binding factor beta / type: protein_or_peptide / ID: 3 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 18.528727 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MPRVVPDQRS KFENEEFFRK LSRECEIKYT GFRDRPHEER QARFQNACRD GRSEIAFVAT GTNLSLQFFP ASWQGEQRQT PSREYVDLE REAGKVYLKA PMILNGVCVI WKGWIDLQRL DGMGCLEFDE ERAQQEDALA QQAFEEARRR TREFEDR

UniProtKB: Core-binding factor subunit beta

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Macromolecule #4: RNA (5'-R(*CP*GP*GP*UP*UP*GP*AP*UP*CP*GP*UP*UP*UP*UP*AP*AP*CP*AP*...

MacromoleculeName: RNA (5'-R(*CP*GP*GP*UP*UP*GP*AP*UP*CP*GP*UP*UP*UP*UP*AP*AP*CP*AP*A)-3')
type: rna / ID: 4 / Number of copies: 2
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 6.369807 KDa
SequenceString:
CGGUUGAUCG UUUUAACAAA

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Macromolecule #5: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 5 / Number of copies: 4 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #6: CHLORIDE ION

MacromoleculeName: CHLORIDE ION / type: ligand / ID: 6 / Number of copies: 2 / Formula: CL
Molecular weightTheoretical: 35.453 Da

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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

BufferpH: 8
Component:
ConcentrationName
25.0 mMTris
150.0 mMsodium chloride
0.5 mMTCEP
Sugar embeddingMaterial: Graphene oxide
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Average electron dose: 37.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.5 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2831190
Startup modelType of model: NONE / Details: ab initio 3D map
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 907456
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.3)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.3)
Final 3D classificationSoftware - Name: RELION (ver. 3.1.3)

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Atomic model buiding 1

Initial model
PDB IDChain

3ir2

chain_id: A, source_name: PDB, initial_model_type: experimental model

6nil

chain_id: B, source_name: PDB, initial_model_type: experimental model

6nil

chain_id: C, source_name: PDB, initial_model_type: experimental model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8h0i:
Cryo-EM structure of APOBEC3G-Vif complex

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