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- PDB-8j62: Cryo-EM structure of APOBEC3G-Vif complex -

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Basic information

Entry
Database: PDB / ID: 8j62
TitleCryo-EM structure of APOBEC3G-Vif complex
Components
  • APOBEC3G
  • Core binding factor beta
  • RNA (5'-R(*CP*GP*GP*UP*UP*GP*AP*UP*UP*GP*UP*UP*UP*UP*AP*AP*CP*AP*A)-3')
  • Viral infectivity factor
KeywordsANTIVIRAL PROTEIN / Human antiviral protein / HIV
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 / osteoblast differentiation / Transcriptional regulation of granulopoiesis / protein polyubiquitination / 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
RNA / RNA (> 10) / Core-binding factor subunit beta
Similarity search - Component
Biological speciesHomo sapiens (human)
Human immunodeficiency virus 1
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.5 Å
AuthorsKouno, T. / Shibata, S. / Hyun, J. / Kim, T.G. / Wolf, M.
Funding support Japan, 1items
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
DepositionApr 24, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jul 19, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: APOBEC3G
C: Viral infectivity factor
D: Core binding factor beta
E: Viral infectivity factor
F: Core binding factor beta
X: RNA (5'-R(*CP*GP*GP*UP*UP*GP*AP*UP*UP*GP*UP*UP*UP*UP*AP*AP*CP*AP*A)-3')
B: APOBEC3G
G: Viral infectivity factor
H: Core binding factor beta
I: Viral infectivity factor
J: Core binding factor beta
Y: RNA (5'-R(*CP*GP*GP*UP*UP*GP*AP*UP*UP*GP*UP*UP*UP*UP*AP*AP*CP*AP*A)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)247,42414
Polymers247,29412
Non-polymers1312
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration, microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein APOBEC3G /


Mass: 43857.266 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#2: Protein
Viral infectivity factor


Mass: 18180.643 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human immunodeficiency virus 1 / Production host: Escherichia coli (E. coli)
#3: Protein
Core binding factor beta


Mass: 18528.727 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli) / References: UniProt: Q13951
#4: RNA chain RNA (5'-R(*CP*GP*GP*UP*UP*GP*AP*UP*UP*GP*UP*UP*UP*UP*AP*AP*CP*AP*A)-3')


Mass: 6370.791 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#5: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn
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

Component
IDNameTypeEntity IDParent-IDSource
1A3G-VC-RNA complexCOMPLEX#1-#40MULTIPLE SOURCES
2APOBEC3G,COre binding factorCOMPLEX#1, #31RECOMBINANT
3Viral infectibity factorCOMPLEX#21RECOMBINANT
4RNACOMPLEX#41SYNTHETIC
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Homo sapiens (human)9606
33Human immunodeficiency virus 111676
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
Buffer component
IDConc.NameBuffer-ID
125 mMTris1
2150 mMNaClSodium chloride1
30.5 mMTCEP1
SpecimenEmbedding applied: YES / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid type: Quantifoil R1.2/1.3
EM embeddingMaterial: Graphene oxide
VitrificationCryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277 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: OTHER
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2400 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 37 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameVersionCategory
7PHENIX1.19.2model fitting
8Coot0.8.9.2model fitting
10RELION3.1.3initial Euler assignment
11RELION3.1.3final Euler assignment
13RELION3.1.33D reconstruction
14PHENIX1.19.2model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 186943 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model buildingPDB-ID: 8H0I

8h0i


Accession code: 8H0I / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00210822
ELECTRON MICROSCOPYf_angle_d0.44914854
ELECTRON MICROSCOPYf_dihedral_angle_d11.4621726
ELECTRON MICROSCOPYf_chiral_restr0.0411594
ELECTRON MICROSCOPYf_plane_restr0.0031728

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