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- EMDB-10044: SIVrcm intasome (Q148/G140S) in complex with bictegravir -

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Basic information

Entry
Database: EMDB / ID: EMD-10044
TitleSIVrcm intasome (Q148/G140S) in complex with bictegravir
Map data
SampleSIVrcm intasome in complex with bictegravir:
Pol Protein / DNA / Pol protein / (nucleic-acidNucleic acid) x 2 / (ligand) x 5
Function / homology
Function and homology information


exoribonuclease H activity / DNA integration / viral genome integration into host DNA / RNA-directed DNA polymerase activity / establishment of integrated proviral latency / RNA-DNA hybrid ribonuclease activity / viral entry into host cell / DNA recombination / aspartic-type endopeptidase activity / DNA binding / zinc ion binding
Integrase, catalytic core / Ribonuclease H superfamily / Retropepsins / Integrase-like, N-terminal / Ribonuclease H-like superfamily / Reverse transcriptase thumb / Reverse transcriptase connection / Integrase, N-terminal zinc-binding domain / Ribonuclease H domain / Peptidase A2A, retrovirus, catalytic ...Integrase, catalytic core / Ribonuclease H superfamily / Retropepsins / Integrase-like, N-terminal / Ribonuclease H-like superfamily / Reverse transcriptase thumb / Reverse transcriptase connection / Integrase, N-terminal zinc-binding domain / Ribonuclease H domain / Peptidase A2A, retrovirus, catalytic / Aspartic peptidase, active site / Integrase, C-terminal, retroviral / Integrase, C-terminal domain superfamily, retroviral / Reverse transcriptase domain / Aspartic peptidase domain superfamily
Pol protein
Biological speciesSimian immunodeficiency virus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.05 Å
AuthorsCherepanov P / Cook N
Funding support United States, United Kingdom, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical SciencesGM082251 United States
The Francis Crick InstituteFC001061 United Kingdom
CitationJournal: Science / Year: 2020
Title: Structural basis of second-generation HIV integrase inhibitor action and viral resistance.
Authors: Nicola J Cook / Wen Li / Dénes Berta / Magd Badaoui / Allison Ballandras-Colas / Andrea Nans / Abhay Kotecha / Edina Rosta / Alan N Engelman / Peter Cherepanov /
Abstract: Despite worldwide prescription, the mechanistic basis for superiority of second-generation HIV integrase (IN) strand transfer inhibitors (INSTIs) is poorly understood. We use single-particle cryo- ...Despite worldwide prescription, the mechanistic basis for superiority of second-generation HIV integrase (IN) strand transfer inhibitors (INSTIs) is poorly understood. We use single-particle cryo-electron microscopy to visualize the mode of action of the advanced INSTIs dolutegravir and bictegravir at near atomic resolution. Q148H/G140S amino acid substitutions in IN that pervade clinical INSTI failure perturb optimal magnesium ion coordination in the enzyme active site. The expanded chemical scaffolds of second-generation compounds mediate interactions with the protein backbone, which are critical for antagonizing Q148H/G140S mutant virus. Our results reveal that binding to magnesium ions underpins a fundamental weakness of the INSTI pharmacophore that is exploited by the virus to engender resistance and provide a structural framework for the development of this important class of anti-HIV/AIDS therapeutics.
Validation ReportPDB-ID: 6rwo

SummaryFull reportAbout validation report
History
DepositionJun 5, 2019-
Header (metadata) releaseJun 12, 2019-
Map releaseFeb 5, 2020-
UpdateFeb 5, 2020-
Current statusFeb 5, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1.6
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 1.6
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6rwo
  • Surface level: 1.6
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic models: PDB-6rwo
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_10044.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.38 Å/pix.
x 360 pix.
= 496.8 Å
1.38 Å/pix.
x 360 pix.
= 496.8 Å
1.38 Å/pix.
x 360 pix.
= 496.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.38 Å
Density
Contour LevelBy AUTHOR: 1.9 / Movie #1: 1.6
Minimum - Maximum-9.949314 - 15.345721
Average (Standard dev.)0.0026487114 (±0.46489707)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 496.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.381.381.38
M x/y/z360360360
origin x/y/z0.0000.0000.000
length x/y/z496.800496.800496.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS360360360
D min/max/mean-9.94915.3460.003

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Supplemental data

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Segmentation: #1

Fileemd_10044_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire SIVrcm intasome in complex with bictegravir

EntireName: SIVrcm intasome in complex with bictegravir / Number of components: 11

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Component #1: protein, SIVrcm intasome in complex with bictegravir

ProteinName: SIVrcm intasome in complex with bictegravir / Recombinant expression: No

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Component #2: protein, Pol Protein

ProteinName: Pol Protein / Recombinant expression: No
SourceSpecies: Simian immunodeficiency virus
Source (engineered)Expression System: Escherichia coli K-12 (bacteria)

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Component #3: protein, DNA

ProteinName: DNA / Recombinant expression: No
SourceSpecies: Simian immunodeficiency virus
Source (engineered)Expression System: synthetic construct (others)

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Component #4: protein, Pol protein

ProteinName: Pol protein / Details: SIVrcm integrase / Number of Copies: 12 / Recombinant expression: No
MassTheoretical: 32.772223 kDa
SourceSpecies: Simian immunodeficiency virus
Source (engineered)Expression System: Escherichia coli K-12 (bacteria)

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Component #5: nucleic-acid, DNA (5'-D(*AP*AP*CP*TP*GP*GP*TP*AP*GP*AP*GP*AP*TP*T...

nucleic acidName: DNA (5'-D(*AP*AP*CP*TP*GP*GP*TP*AP*GP*AP*GP*AP*TP*TP*TP*TP*TP*CP*TP*TP*AP*GP*C)-3')
Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DA)(DA)(DC)(DT)(DG)(DG)(DT)(DA)(DG)(DA) (DG)(DA)(DT)(DT)(DT)(DT)(DT)(DC)(DT)(DT) (DA)(DG)(DC)(DC)(DT)(DT)(DC)(DT)(DA)(DG) (DA)(DA)(DC)
MassTheoretical: 10.134541 kDa
SourceSpecies: Simian immunodeficiency virus

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Component #6: nucleic-acid, DNA (5'-D(P*GP*CP*TP*AP*AP*GP*AP*AP*AP*AP*AP*TP*CP*...

nucleic acidName: DNA (5'-D(P*GP*CP*TP*AP*AP*GP*AP*AP*AP*AP*AP*TP*CP*TP*CP*TP*AP*CP*CP*A)-3')
Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DG)(DT)(DT)(DC)(DT)(DA)(DG)(DA)(DA)(DG) (DG)(DC)(DT)(DA)(DA)(DG)(DA)(DA)(DA)(DA) (DA)(DT)(DC)(DT)(DC)(DT)(DA)(DC)(DC)(DA)
MassTheoretical: 9.223995 kDa
SourceSpecies: Simian immunodeficiency virus

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Component #7: ligand, ZINC ION

LigandName: ZINC ION / Number of Copies: 8 / Recombinant expression: No
MassTheoretical: 6.540905 MDa

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Component #8: ligand, CHLORIDE ION

LigandName: CHLORIDE IONChloride / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 3.545305 MDa

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Component #9: ligand, Bictegravir

LigandName: Bictegravir / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.44938 kDa

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Component #10: ligand, MAGNESIUM ION

LigandName: MAGNESIUM ION / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 2.430505 MDa

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Component #11: ligand, water

LigandName: water / Number of Copies: 6 / Recombinant expression: No
MassTheoretical: 1.801505 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionpH: 7
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 295 K / Humidity: 95 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 50.4 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1600.0 - 3500.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 6618

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 217635
3D reconstructionAlgorithm: FOURIER SPACE / Software: cryoSPARC / Resolution: 3.05 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Modeling #1Refinement protocol: rigid body / Refinement space: REAL
Input PDB model: 2B4J, 1EX4, 1K6Y
Output model

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