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- EMDB-13076: STLV-1 intasome:B56 in complex with the strand-transfer inhibitor... -

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Basic information

Entry
Database: EMDB / ID: EMD-13076
TitleSTLV-1 intasome:B56 in complex with the strand-transfer inhibitor raltegravir
Map dataDeemEMenhancer highest target
Sample
  • Complex: Complex of STLV-1 MarB43 integrase with nascent viral DNA, the human PP2A B56 subunit and the inhibitor raltegravir
    • Protein or peptide: Integrase
    • Protein or peptide: Isoform 3 of PC4 and SFRS1-interacting protein,Isoform Gamma-2 of Serine/threonine-protein phosphatase 2A 56 kDa regulatory subunit gamma isoform
    • DNA: DNA (5'-D(*AP*CP*TP*GP*TP*GP*TP*TP*TP*GP*GP*CP*GP*CP*TP*TP*CP*TP*CP*TP*C)-3')
    • DNA: DNA (5'-D(*GP*AP*GP*AP*GP*AP*AP*GP*CP*GP*CP*CP*AP*AP*AP*CP*AP*CP*A)-3')
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
  • Ligand: N-(4-fluorobenzyl)-5-hydroxy-1-methyl-2-(1-methyl-1-{[(5-methyl-1,3,4-oxadiazol-2-yl)carbonyl]amino}ethyl)-6-oxo-1,6-di hydropyrimidine-4-carboxamide
  • Ligand: water
Function / homology
Function and homology information


protein phosphatase type 2A complex / protein phosphatase regulator activity / APC truncation mutants have impaired AXIN binding / AXIN missense mutants destabilize the destruction complex / Truncations of AMER1 destabilize the destruction complex / Beta-catenin phosphorylation cascade / Signaling by GSK3beta mutants / CTNNB1 S33 mutants aren't phosphorylated / CTNNB1 S37 mutants aren't phosphorylated / CTNNB1 S45 mutants aren't phosphorylated ...protein phosphatase type 2A complex / protein phosphatase regulator activity / APC truncation mutants have impaired AXIN binding / AXIN missense mutants destabilize the destruction complex / Truncations of AMER1 destabilize the destruction complex / Beta-catenin phosphorylation cascade / Signaling by GSK3beta mutants / CTNNB1 S33 mutants aren't phosphorylated / CTNNB1 S37 mutants aren't phosphorylated / CTNNB1 S45 mutants aren't phosphorylated / CTNNB1 T41 mutants aren't phosphorylated / supercoiled DNA binding / Disassembly of the destruction complex and recruitment of AXIN to the membrane / Integration of viral DNA into host genomic DNA / Autointegration results in viral DNA circles / CTLA4 inhibitory signaling / Platelet sensitization by LDL / Formation of WDR5-containing histone-modifying complexes / 2-LTR circle formation / Vpr-mediated nuclear import of PICs / protein phosphatase activator activity / Integration of provirus / APOBEC3G mediated resistance to HIV-1 infection / mRNA 5'-splice site recognition / chromosome, centromeric region / intrinsic apoptotic signaling pathway in response to DNA damage by p53 class mediator / heterochromatin / Amplification of signal from unattached kinetochores via a MAD2 inhibitory signal / DNA damage response, signal transduction by p53 class mediator resulting in cell cycle arrest / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / Resolution of Sister Chromatid Cohesion / nuclear periphery / RHO GTPases Activate Formins / RAF activation / euchromatin / Degradation of beta-catenin by the destruction complex / DNA integration / Negative regulation of MAPK pathway / viral genome integration into host DNA / Separation of Sister Chromatids / establishment of integrated proviral latency / RNA-directed DNA polymerase activity / RNA-DNA hybrid ribonuclease activity / Regulation of TP53 Degradation / response to heat / PI5P, PP2A and IER3 Regulate PI3K/AKT Signaling / proteasome-mediated ubiquitin-dependent protein catabolic process / DNA-binding transcription factor binding / DNA recombination / response to oxidative stress / nucleic acid binding / transcription coactivator activity / chromatin remodeling / negative regulation of cell population proliferation / chromatin binding / Golgi apparatus / signal transduction / positive regulation of transcription by RNA polymerase II / RNA binding / zinc ion binding / nucleoplasm / nucleus / cytosol
Similarity search - Function
Protein phosphatase 2A, regulatory B subunit, B56 / Protein phosphatase 2A regulatory B subunit (B56 family) / Lens epithelium-derived growth factor, integrase-binding domain / HIV integrase-binding domain superfamily / Lens epithelium-derived growth factor (LEDGF) / TFIIS/LEDGF domain superfamily / domain with conserved PWWP motif / PWWP domain / PWWP domain profile. / PWWP domain ...Protein phosphatase 2A, regulatory B subunit, B56 / Protein phosphatase 2A regulatory B subunit (B56 family) / Lens epithelium-derived growth factor, integrase-binding domain / HIV integrase-binding domain superfamily / Lens epithelium-derived growth factor (LEDGF) / TFIIS/LEDGF domain superfamily / domain with conserved PWWP motif / PWWP domain / PWWP domain profile. / PWWP domain / Integrase Zinc binding domain / Integrase DNA binding domain / Integrase, N-terminal zinc-binding domain / Integrase, C-terminal, retroviral / Integrase DNA binding domain profile. / RNase H / Integrase core domain / Integrase, catalytic core / Integrase catalytic domain profile. / Ribonuclease H domain / RNase H type-1 domain profile. / Reverse transcriptase (RNA-dependent DNA polymerase) / Reverse transcriptase domain / Reverse transcriptase (RT) catalytic domain profile. / Armadillo-like helical / Armadillo-type fold / Ribonuclease H superfamily / Ribonuclease H-like superfamily / Reverse transcriptase/Diguanylate cyclase domain / DNA/RNA polymerase superfamily
Similarity search - Domain/homology
PC4 and SFRS1-interacting protein / Serine/threonine-protein phosphatase 2A 56 kDa regulatory subunit gamma isoform / Pol protein
Similarity search - Component
Biological speciesSimian T-lymphotropic virus 1 / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsBarski MS / Ballandras-Colas A / Cronin NB / Pye VE / Cherepanov P / Maertens GN
Funding support United Kingdom, United States, 4 items
OrganizationGrant numberCountry
Wellcome Trust107005/Z/15Z United Kingdom
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)GM082251 United States
The Francis Crick InstituteFC001061 United Kingdom
Wellcome Trust206175/Z/17/Z United Kingdom
CitationJournal: Nat Commun / Year: 2021
Title: Structural basis for the inhibition of HTLV-1 integration inferred from cryo-EM deltaretroviral intasome structures.
Authors: Michal S Barski / Teresa Vanzo / Xue Zhi Zhao / Steven J Smith / Allison Ballandras-Colas / Nora B Cronin / Valerie E Pye / Stephen H Hughes / Terrence R Burke / Peter Cherepanov / Goedele N Maertens /
Abstract: Between 10 and 20 million people worldwide are infected with the human T-cell lymphotropic virus type 1 (HTLV-1). Despite causing life-threatening pathologies there is no therapeutic regimen for this ...Between 10 and 20 million people worldwide are infected with the human T-cell lymphotropic virus type 1 (HTLV-1). Despite causing life-threatening pathologies there is no therapeutic regimen for this deltaretrovirus. Here, we screened a library of integrase strand transfer inhibitor (INSTI) candidates built around several chemical scaffolds to determine their effectiveness in limiting HTLV-1 infection. Naphthyridines with substituents in position 6 emerged as the most potent compounds against HTLV-1, with XZ450 having highest efficacy in vitro. Using single-particle cryo-electron microscopy we visualised XZ450 as well as the clinical HIV-1 INSTIs raltegravir and bictegravir bound to the active site of the deltaretroviral intasome. The structures reveal subtle differences in the coordination environment of the Mg ion pair involved in the interaction with the INSTIs. Our results elucidate the binding of INSTIs to the HTLV-1 intasome and support their use for pre-exposure prophylaxis and possibly future treatment of HTLV-1 infection.
History
DepositionJun 11, 2021-
Header (metadata) releaseAug 18, 2021-
Map releaseAug 18, 2021-
UpdateOct 6, 2021-
Current statusOct 6, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7oug
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_13076.png

Downloads & links

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Map

FileDownload / File: emd_13076.map.gz / Format: CCP4 / Size: 107.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDeemEMenhancer highest target
Voxel sizeX=Y=Z: 1.1 Å
Density
Contour LevelBy AUTHOR: 0.0187 / Movie #1: 0.05
Minimum - Maximum-0.024642875 - 1.9791597
Average (Standard dev.)0.00090026844 (±0.021383742)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions304304304
Spacing304304304
CellA=B=C: 334.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.11.11.1
M x/y/z304304304
origin x/y/z0.0000.0000.000
length x/y/z334.400334.400334.400
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ540540540
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS304304304
D min/max/mean-0.0251.9790.001

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Supplemental data

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Additional map: denmod map which was used for model real space refinement

Fileemd_13076_additional_1.map
Annotationdenmod map which was used for model real space refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 2

Fileemd_13076_half_map_1.map
Annotationhalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 1

Fileemd_13076_half_map_2.map
Annotationhalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Complex of STLV-1 MarB43 integrase with nascent viral DNA, the hu...

EntireName: Complex of STLV-1 MarB43 integrase with nascent viral DNA, the human PP2A B56 subunit and the inhibitor raltegravir
Components
  • Complex: Complex of STLV-1 MarB43 integrase with nascent viral DNA, the human PP2A B56 subunit and the inhibitor raltegravir
    • Protein or peptide: Integrase
    • Protein or peptide: Isoform 3 of PC4 and SFRS1-interacting protein,Isoform Gamma-2 of Serine/threonine-protein phosphatase 2A 56 kDa regulatory subunit gamma isoform
    • DNA: DNA (5'-D(*AP*CP*TP*GP*TP*GP*TP*TP*TP*GP*GP*CP*GP*CP*TP*TP*CP*TP*CP*TP*C)-3')
    • DNA: DNA (5'-D(*GP*AP*GP*AP*GP*AP*AP*GP*CP*GP*CP*CP*AP*AP*AP*CP*AP*CP*A)-3')
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
  • Ligand: N-(4-fluorobenzyl)-5-hydroxy-1-methyl-2-(1-methyl-1-{[(5-methyl-1,3,4-oxadiazol-2-yl)carbonyl]amino}ethyl)-6-oxo-1,6-di hydropyrimidine-4-carboxamide
  • Ligand: water

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Supramolecule #1: Complex of STLV-1 MarB43 integrase with nascent viral DNA, the hu...

SupramoleculeName: Complex of STLV-1 MarB43 integrase with nascent viral DNA, the human PP2A B56 subunit and the inhibitor raltegravir
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Details: Sample composition and source have been described in "macromolecules"
Molecular weightTheoretical: 331.1 KDa

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Macromolecule #1: Integrase

MacromoleculeName: Integrase / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Simian T-lymphotropic virus 1
Molecular weightTheoretical: 33.943539 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: GPEFQLSPAK LHSFTHCGQA ALTLHGATTT EALNILHSCH ACRKNNPQHQ MPRGHIRRGL LPNHIWQGDI THFKYKNTLY RLHVWVDTF SGSVSATHKK RETSSEAISS LLHAIAHLGR PSHINTDNGP AYASQEFQHA CTSLAIRHTT HIPYNPTSSG L VERTNGIL ...String:
GPEFQLSPAK LHSFTHCGQA ALTLHGATTT EALNILHSCH ACRKNNPQHQ MPRGHIRRGL LPNHIWQGDI THFKYKNTLY RLHVWVDTF SGSVSATHKK RETSSEAISS LLHAIAHLGR PSHINTDNGP AYASQEFQHA CTSLAIRHTT HIPYNPTSSG L VERTNGIL KTLLYKYFSD NPNLPMDNAL SVALWTINHL NVLTHCQKTR WQLHHSPRLP PIPEEKPVTT SKTHWYYFKI PG LNSRQWK GPQRALQEAA GAALIPVSDT AAQWIPWKLL KRAVCPRLAG DTADPKERDH QHHG

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Macromolecule #2: Isoform 3 of PC4 and SFRS1-interacting protein,Isoform Gamma-2 of...

MacromoleculeName: Isoform 3 of PC4 and SFRS1-interacting protein,Isoform Gamma-2 of Serine/threonine-protein phosphatase 2A 56 kDa regulatory subunit gamma isoform
type: protein_or_peptide / ID: 2
Details: fusion construct containing human LEDGF (residues 1-324 thus without the IBD domain) (gene PSIP1; O75475)) and human B56gammma (residues 11-380) regulatory subunit of PP2A (gene PPP2R5C) (no ...Details: fusion construct containing human LEDGF (residues 1-324 thus without the IBD domain) (gene PSIP1; O75475)) and human B56gammma (residues 11-380) regulatory subunit of PP2A (gene PPP2R5C) (no space for these details below).,fusion construct containing human LEDGF (residues 1-324 thus without the IBD domain) (gene PSIP1; O75475)) and human B56gammma (residues 11-380) regulatory subunit of PP2A (gene PPP2R5C) (no space for these details below).
Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 80.39468 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SMTRDFKPGD LIFAKMKGYP HWPARVDEVP DGAVKPPTNK LPIFFFGTHE TAFLGPKDIF PYSENKEKYG KPNKRKGFNE GLWEIDNNP KVKFSSQQAA TKQSNASSDV EVEEKETSVS KEDTDHEEKA SNEDVTKAVD ITTPKAARRG RKRKAEKQVE T EEAGVVTT ...String:
SMTRDFKPGD LIFAKMKGYP HWPARVDEVP DGAVKPPTNK LPIFFFGTHE TAFLGPKDIF PYSENKEKYG KPNKRKGFNE GLWEIDNNP KVKFSSQQAA TKQSNASSDV EVEEKETSVS KEDTDHEEKA SNEDVTKAVD ITTPKAARRG RKRKAEKQVE T EEAGVVTT ATASVNLKVS PKRGRPAATE VKIPKPRGRP KMVKQPCPSE SDIITEEDKS KKKGQEEKQP KKQPKKDEEG QK EEDKPRK EPDKKEGKKE VESKRKNLAK TGVTSTSDSE EEGDDQEGEK KRKGGRNFQT AHRRNMLKGQ HEKEAADRKR KQE EQMETE FMVVDAANSN GPFQPVVLLH IRDVPPADQE KLFIQKLRQC CVLFDFVSDP LSDLKWKEVK RAALSEMVEY ITHN RNVIT EPIYPEVVHM FAVNMFRTLP PSSNPTGAEF DPEEDEPTLE AAWPHLQLVY EFFLRFLESP DFQPNIAKKY IDQKF VLQL LELFDSEDPR ERDFLKTTLH RIYGKFLGLR AYIRKQINNI FYRFIYETEH HNGIAELLEI LGSIINGFAL PLKEEH KIF LLKVLLPLHK VKSLSVYHPQ LAYCVVQFLE KDSTLTEPVV MALLKYWPKT HSPKEVMFLN ELEEILDVIE PSEFVKI ME PLFRQLAKCV SSPHFQVAER ALYYWNNEYI MSLISDNAAK ILPIMFPSLY RNSKT

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Macromolecule #3: DNA (5'-D(*AP*CP*TP*GP*TP*GP*TP*TP*TP*GP*GP*CP*GP*CP*TP*TP*CP*TP*...

MacromoleculeName: DNA (5'-D(*AP*CP*TP*GP*TP*GP*TP*TP*TP*GP*GP*CP*GP*CP*TP*TP*CP*TP*CP*TP*C)-3')
type: dna / ID: 3 / Number of copies: 2 / Classification: DNA
Source (natural)Organism: Simian T-lymphotropic virus 1
Molecular weightTheoretical: 9.221909 KDa
SequenceString:
(DA)(DC)(DT)(DG)(DT)(DG)(DT)(DT)(DT)(DG) (DG)(DC)(DG)(DC)(DT)(DT)(DC)(DT)(DC)(DT) (DC)(DC)(DC)(DG)(DG)(DA)(DG)(DA)(DG) (DA)

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Macromolecule #4: DNA (5'-D(*GP*AP*GP*AP*GP*AP*AP*GP*CP*GP*CP*CP*AP*AP*AP*CP*AP*CP*...

MacromoleculeName: DNA (5'-D(*GP*AP*GP*AP*GP*AP*AP*GP*CP*GP*CP*CP*AP*AP*AP*CP*AP*CP*A)-3')
type: dna / ID: 4 / Number of copies: 2 / Classification: DNA
Source (natural)Organism: Simian T-lymphotropic virus 1
Molecular weightTheoretical: 8.59356 KDa
SequenceString:
(DT)(DC)(DT)(DC)(DT)(DC)(DC)(DG)(DG)(DG) (DA)(DG)(DA)(DG)(DA)(DA)(DG)(DC)(DG)(DC) (DC)(DA)(DA)(DA)(DC)(DA)(DC)(DA)

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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: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 6 / Number of copies: 4 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #7: N-(4-fluorobenzyl)-5-hydroxy-1-methyl-2-(1-methyl-1-{[(5-methyl-1...

MacromoleculeName: N-(4-fluorobenzyl)-5-hydroxy-1-methyl-2-(1-methyl-1-{[(5-methyl-1,3,4-oxadiazol-2-yl)carbonyl]amino}ethyl)-6-oxo-1,6-di hydropyrimidine-4-carboxamide
type: ligand / ID: 7 / Number of copies: 2 / Formula: RLT
Molecular weightTheoretical: 444.416 Da

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Macromolecule #8: water

MacromoleculeName: water / type: ligand / ID: 8 / Number of copies: 8 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / Water

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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

Concentration0.76 mg/mL
BufferpH: 6
Component:
ConcentrationNameFormula
20.0 mMBis tris propane
0.3 MSodium chlorideNaClSodium chloride
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE
Details: Glow-discharged for 4 min at 45 mA on an Emitech K100X instrument (Electron Microscopy Sciences) and covered with a layer of graphene oxide (Sigma-Aldrich, catalogue #763705) immediately before being used.
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 295.15 K / Instrument: FEI VITROBOT MARK IV
DetailsComplex was isolated by size exclusion chromatography

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Detector mode: COUNTING / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2840121
CTF correctionSoftware - Name: Gctf (ver. 1.06)
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: cryoSPARC
Final 3D classificationSoftware - Name: RELION (ver. 3.1)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 3.1)
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 111051
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6z2y
PDB Unreleased entry


Details: 6Z2Y was fitted into the cryoEM map using Chimera. The model was adjusted to fit the map; metal ions and drug docked into the map manually using Coot. The final model was subjected to Phenix. ...Details: 6Z2Y was fitted into the cryoEM map using Chimera. The model was adjusted to fit the map; metal ions and drug docked into the map manually using Coot. The final model was subjected to Phenix.real_space_refine using C2 NCS, secondary structure and metal ion coordination restraints.
Details6Z2Y was fitted into the cryoEM map using Chimera. The model was adjusted to fit the map; metal ions and drug docked into the map manually using Coot. The final model was subjected to Phenix.real_space_refine using C2 NCS, secondary structure and metal ion coordination restraints.
RefinementProtocol: RIGID BODY FIT
Output model

PDB-7oug:
STLV-1 intasome:B56 in complex with the strand-transfer inhibitor raltegravir

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