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- PDB-6puw: Structure of HIV cleaved synaptic complex (CSC) intasome bound wi... -

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Basic information

Entry
Database: PDB / ID: 6puw
TitleStructure of HIV cleaved synaptic complex (CSC) intasome bound with magnesium and Bictegravir (BIC)
Components
  • Chimeric Sso7d and HIV-1 integrase
  • viral DNA non-transferred strand
  • viral DNA transferred strand
KeywordsVIRAL PROTEIN/DNA / integrase / intasome / transposition / VIRAL PROTEIN / VIRAL PROTEIN-DNA complex
Function / homology
Function and homology information


RNA endonuclease activity / DNA integration / viral genome integration into host DNA / establishment of integrated proviral latency / RNA-directed DNA polymerase activity / endonuclease activity / DNA recombination / symbiont entry into host cell / DNA binding / zinc ion binding / cytoplasm
Similarity search - Function
Integrase, C-terminal domain superfamily, retroviral / DNA-binding 7kDa protein / 7kD DNA-binding domain / Chromo-like domain superfamily / Integrase Zinc binding domain / Zinc finger integrase-type profile. / Integrase-like, N-terminal / Integrase DNA binding domain / Integrase, C-terminal domain superfamily, retroviral / Integrase, N-terminal zinc-binding domain ...Integrase, C-terminal domain superfamily, retroviral / DNA-binding 7kDa protein / 7kD DNA-binding domain / Chromo-like domain superfamily / Integrase Zinc binding domain / Zinc finger integrase-type profile. / Integrase-like, N-terminal / Integrase DNA binding domain / Integrase, C-terminal domain superfamily, retroviral / Integrase, N-terminal zinc-binding domain / Integrase, C-terminal, retroviral / Integrase DNA binding domain profile. / Integrase core domain / Integrase, catalytic core / Integrase catalytic domain profile. / SH3 type barrels. / Ribonuclease H superfamily / Ribonuclease H-like superfamily / Roll / Mainly Beta
Similarity search - Domain/homology
Bictegravir / DNA / DNA (> 10) / DNA-binding protein 7d / Integrase
Similarity search - Component
Biological speciesSaccharolobus solfataricus (archaea)
Human immunodeficiency virus 1
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsLyumkis, D. / Jozwik, I.K. / Passos, D.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01 AI136680 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM069832 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01 AI146017 United States
CitationJournal: Science / Year: 2020
Title: Structural basis for strand-transfer inhibitor binding to HIV intasomes.
Authors: Dario Oliveira Passos / Min Li / Ilona K Jóźwik / Xue Zhi Zhao / Diogo Santos-Martins / Renbin Yang / Steven J Smith / Youngmin Jeon / Stefano Forli / Stephen H Hughes / Terrence R Burke / ...Authors: Dario Oliveira Passos / Min Li / Ilona K Jóźwik / Xue Zhi Zhao / Diogo Santos-Martins / Renbin Yang / Steven J Smith / Youngmin Jeon / Stefano Forli / Stephen H Hughes / Terrence R Burke / Robert Craigie / Dmitry Lyumkis /
Abstract: The HIV intasome is a large nucleoprotein assembly that mediates the integration of a DNA copy of the viral genome into host chromatin. Intasomes are targeted by the latest generation of ...The HIV intasome is a large nucleoprotein assembly that mediates the integration of a DNA copy of the viral genome into host chromatin. Intasomes are targeted by the latest generation of antiretroviral drugs, integrase strand-transfer inhibitors (INSTIs). Challenges associated with lentiviral intasome biochemistry have hindered high-resolution structural studies of how INSTIs bind to their native drug target. Here, we present high-resolution cryo-electron microscopy structures of HIV intasomes bound to the latest generation of INSTIs. These structures highlight how small changes in the integrase active site can have notable implications for drug binding and design and provide mechanistic insights into why a leading INSTI retains efficacy against a broad spectrum of drug-resistant variants. The data have implications for expanding effective treatments available for HIV-infected individuals.
History
DepositionJul 18, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Feb 12, 2020Provider: repository / Type: Initial release
Revision 1.1Feb 26, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Mar 20, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model / pdbx_struct_oper_list
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type / _pdbx_struct_oper_list.name / _pdbx_struct_oper_list.symmetry_operation / _pdbx_struct_oper_list.type

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Structure visualization

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  • Biological unit as complete point assembly
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  • Deposited structure unit
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  • Simplified surface model + fitted atomic model
  • EMDB-20483
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Structure viewerMolecule:
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Assembly

Deposited unit
A: Chimeric Sso7d and HIV-1 integrase
B: Chimeric Sso7d and HIV-1 integrase
C: Chimeric Sso7d and HIV-1 integrase
D: Chimeric Sso7d and HIV-1 integrase
E: viral DNA non-transferred strand
F: viral DNA transferred strand
hetero molecules


Theoretical massNumber of molelcules
Total (without water)185,87511
Polymers185,2466
Non-polymers6295
Water2,810156
1
A: Chimeric Sso7d and HIV-1 integrase
B: Chimeric Sso7d and HIV-1 integrase
C: Chimeric Sso7d and HIV-1 integrase
D: Chimeric Sso7d and HIV-1 integrase
E: viral DNA non-transferred strand
F: viral DNA transferred strand
hetero molecules

A: Chimeric Sso7d and HIV-1 integrase
B: Chimeric Sso7d and HIV-1 integrase
C: Chimeric Sso7d and HIV-1 integrase
D: Chimeric Sso7d and HIV-1 integrase
E: viral DNA non-transferred strand
F: viral DNA transferred strand
hetero molecules


Theoretical massNumber of molelcules
Total (without water)371,75022
Polymers370,49312
Non-polymers1,25810
Water21612
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation1
2


  • Idetical with deposited unit
  • point asymmetric unit
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3


  • Idetical with deposited unit in distinct coordinate
  • point asymmetric unit, std point frame
TypeNameSymmetry operationNumber
transform to point frame1
SymmetryPoint symmetry: (Schoenflies symbol: C2 (2 fold cyclic))

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Components

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Protein , 1 types, 4 molecules ABCD

#1: Protein
Chimeric Sso7d and HIV-1 integrase / 7 kDa DNA-binding protein d / Sso7d


Mass: 42321.258 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharolobus solfataricus (strain ATCC 35092 / DSM 1617 / JCM 11322 / P2) (archaea), (gene. exp.) Human immunodeficiency virus 1
Strain: ATCC 35092 / DSM 1617 / JCM 11322 / P2 / Gene: sso7d, sso7d-1, SSO10610 / Production host: Escherichia coli (E. coli) / References: UniProt: P39476, UniProt: Q76353

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DNA chain , 2 types, 2 molecules EF

#2: DNA chain viral DNA non-transferred strand


Mass: 8188.271 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Human immunodeficiency virus 1
#3: DNA chain viral DNA transferred strand


Mass: 7773.023 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Human immunodeficiency virus 1

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Non-polymers , 4 types, 161 molecules

#4: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#5: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn
#6: Chemical ChemComp-KLQ / Bictegravir / Bictegravir


Mass: 449.380 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C21H18F3N3O5 / Feature type: SUBJECT OF INVESTIGATION / Comment: inhibitor*YM
#7: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 156 / Source method: isolated from a natural source / Formula: H2O

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Details

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: Assembly of HIV integrase and viral DNA / Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT
Molecular weightValue: 0.4 MDa / Experimental value: NO
Source (natural)Organism: hiv (virus)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 6.2
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMBis-TrisBis-tris methane1
2550 mMsodium chlorideNaClSodium chloride1
35 mMmagnesium chlorideMgCl1
40.5 mMTCEP1
55 % (w/v)glycerol1
SpecimenConc.: 0.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 400 divisions/in. / Grid type: UltrAuFoil
VitrificationInstrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 80 %

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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 FIELDBright-field microscopy / Nominal magnification: 37000 X / Calibrated magnification: 63291 X / Nominal defocus max: 3000 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 45 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 2188
Image scansWidth: 3838 / Height: 3710 / Movie frames/image: 80 / Used frames/image: 4-80

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Processing

SoftwareName: PHENIX / Version: (1.16_3549: ???) / Classification: refinement
EM software
IDNameVersionCategory
2Leginonimage acquisition
4WarpCTF correction
7Cootmodel fitting
9PHENIX1.16model refinement
10cisTEM1initial Euler assignment
11cisTEM1.0 betafinal Euler assignment
12cisTEM1.0 betaclassification
13cisTEM1.0 beta3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 340266
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 146022 / Algorithm: FOURIER SPACE / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL / Target criteria: Correlation coefficient
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
13L3U

3l3u

13L3U1PDBexperimental model
21K6Y

1k6y

11K6Y2PDBexperimental model
35U1C

5u1c

15U1C3PDBexperimental model
RefinementResolution: 2.9→214.508 Å / SU ML: 1.18 / Phase error: 60.17 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.5345 2014 0.09 %
Rwork0.5426 --
obs0.5426 2164272 99.96 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0055579
ELECTRON MICROSCOPYf_angle_d0.5827683
ELECTRON MICROSCOPYf_dihedral_angle_d22.2562142
ELECTRON MICROSCOPYf_chiral_restr0.043854
ELECTRON MICROSCOPYf_plane_restr0.003837
LS refinement shellResolution: 2.7002→2.763 Å
RfactorNum. reflection% reflection
Rfree1.2589 144 -
Rwork1.4219 197541 -
obs--100 %

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