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- PDB-9nlp: HIV-1 Reverse Transcriptase with New Non-Nucleoside Reverse Trans... -

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Basic information

Entry
Database: PDB / ID: 9nlp
TitleHIV-1 Reverse Transcriptase with New Non-Nucleoside Reverse Transcriptase Inhibitor 12126065
Components
  • Reverse transcriptase p51 subunit
  • Reverse transcriptase p66 subunit
KeywordsVIRAL PROTEIN / Reverse transcriptase
Function / homology
Function and homology information


HIV-1 retropepsin / symbiont-mediated activation of host apoptosis / retroviral ribonuclease H / exoribonuclease H / exoribonuclease H activity / host multivesicular body / DNA integration / viral genome integration into host DNA / RNA-directed DNA polymerase / establishment of integrated proviral latency ...HIV-1 retropepsin / symbiont-mediated activation of host apoptosis / retroviral ribonuclease H / exoribonuclease H / exoribonuclease H activity / host multivesicular body / DNA integration / viral genome integration into host DNA / RNA-directed DNA polymerase / establishment of integrated proviral latency / telomerase activity / viral penetration into host nucleus / RNA stem-loop binding / RNA-DNA hybrid ribonuclease activity / Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases / host cell / viral nucleocapsid / DNA recombination / DNA-directed DNA polymerase / aspartic-type endopeptidase activity / Hydrolases; Acting on ester bonds / DNA-directed DNA polymerase activity / symbiont-mediated suppression of host gene expression / symbiont entry into host cell / viral translational frameshifting / lipid binding / host cell nucleus / host cell plasma membrane / virion membrane / structural molecule activity / proteolysis / DNA binding / zinc ion binding / membrane
Similarity search - Function
Reverse transcriptase connection / Reverse transcriptase connection domain / Reverse transcriptase thumb / Reverse transcriptase thumb domain / 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 ...Reverse transcriptase connection / Reverse transcriptase connection domain / Reverse transcriptase thumb / Reverse transcriptase thumb domain / 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. / Immunodeficiency lentiviral matrix, N-terminal / gag gene protein p17 (matrix protein) / RNase H / Integrase core domain / Integrase, catalytic core / Integrase catalytic domain profile. / Retropepsin-like catalytic domain / Matrix protein, lentiviral and alpha-retroviral, N-terminal / Retroviral nucleocapsid Gag protein p24, C-terminal domain / Gag protein p24 C-terminal domain / RNase H type-1 domain profile. / Ribonuclease H domain / Retropepsins / Retroviral aspartyl protease / Aspartyl protease, retroviral-type family profile. / Peptidase A2A, retrovirus, catalytic / Reverse transcriptase domain / Reverse transcriptase (RNA-dependent DNA polymerase) / Reverse transcriptase (RT) catalytic domain profile. / Retrovirus capsid, C-terminal / Retroviral matrix protein / Retrovirus capsid, N-terminal / zinc finger / Zinc knuckle / Zinc finger, CCHC-type superfamily / Zinc finger, CCHC-type / Zinc finger CCHC-type profile. / Aspartic peptidase, active site / Eukaryotic and viral aspartyl proteases active site. / Aspartic peptidase domain superfamily / Ribonuclease H superfamily / Ribonuclease H-like superfamily / Reverse transcriptase/Diguanylate cyclase domain / DNA/RNA polymerase superfamily
Similarity search - Domain/homology
: / Gag-Pol polyprotein
Similarity search - Component
Biological speciesHIV-1 06TG.HT008 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.53 Å
AuthorsYoung, M.A. / Lane, T.R. / Raman, R. / Nelson, J.A.E. / Riabova, O. / Kazakova, E. / Monakhova, N. / Tsedilin, A. / Rees, S.D. / Quinnell, D. ...Young, M.A. / Lane, T.R. / Raman, R. / Nelson, J.A.E. / Riabova, O. / Kazakova, E. / Monakhova, N. / Tsedilin, A. / Rees, S.D. / Quinnell, D. / Chang, G. / Ekins, S.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: ACS Infect Dis / Year: 2025
Title: Cryo-EM Structure of HIV-1 Reverse Transcriptase with -Phenyl-1-(phenylsulfonyl)-1-1,2,4-triazol-3-amine: A New HIV-1 Non-nucleoside Inhibitor.
Authors: Megan A Young / Thomas R Lane / Renuka Raman / Julie A E Nelson / Olga Riabova / Elena Kazakova / Natalia Monakhova / Andrey Tsedilin / Steven D Rees / Daniel Quinnell / Vadim Makarov / ...Authors: Megan A Young / Thomas R Lane / Renuka Raman / Julie A E Nelson / Olga Riabova / Elena Kazakova / Natalia Monakhova / Andrey Tsedilin / Steven D Rees / Daniel Quinnell / Vadim Makarov / Geoffrey Chang / Sean Ekins /
Abstract: The use of highly active antiretroviral therapy (HAART) has made the human immunodeficiency virus (HIV) a chronic disease rather than a terminal disease. Non-nucleoside reverse transcriptase ...The use of highly active antiretroviral therapy (HAART) has made the human immunodeficiency virus (HIV) a chronic disease rather than a terminal disease. Non-nucleoside reverse transcriptase inhibitors (NNRTIs) are an important component of HAART, although we are seeing clinically relevant drug-resistant mutants such that there is a need to develop new molecules. We recently identified a new class of -phenyl-1-(phenylsulfonyl)-1-1,2,4-triazol-3-amine HIV-1 NNRTI, with one known as compound 12126065, with sub nanomolar (nM) potency in TZM-bl cells (HeLa cells expressing CD4, CCR5, and CXCR4) with no in vivo acute or subacute toxicity. We now describe the cryo-EM structure of this molecule (resolution of 3.53 Å) and compare it to analogues and other known NNRTIs. We also describe the synthesis and activity of five additional analogues of this class of compounds, some of which have promising activity against a K103N/Y181C (A17) double mutant, which will enable the design of future molecules.
History
DepositionMar 3, 2025Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 7, 2025Provider: repository / Type: Initial release
Revision 1.1May 21, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Reverse transcriptase p66 subunit
B: Reverse transcriptase p51 subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)117,8283
Polymers117,3502
Non-polymers4781
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Reverse transcriptase p66 subunit / Exoribonuclease H / p66 RT


Mass: 65884.500 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) HIV-1 06TG.HT008 (virus) / Gene: gag-pol / Production host: Escherichia coli (E. coli) / References: UniProt: P12497, RNA-directed DNA polymerase
#2: Protein Reverse transcriptase p51 subunit


Mass: 51465.234 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) HIV-1 06TG.HT008 (virus) / Gene: gag-pol / Production host: Escherichia coli (E. coli) / References: UniProt: P12497, RNA-directed DNA polymerase
#3: Chemical ChemComp-A1BYY / 4-({5-amino-1-[6-(2-cyanoethyl)naphthalene-1-sulfonyl]-1H-1,2,4-triazol-3-yl}amino)-2-chlorobenzonitrile


Mass: 477.926 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C22H16ClN7O2S / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY
Has protein modificationN

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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: HIV-1 reverse transcriptase with novel inhibitor / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightValue: 1.17 MDa / Experimental value: YES
Source (natural)Organism: HIV-1 06TG.HT008 (virus)
Source (recombinant)Organism: Escherichia coli (E. coli)
Details of virusEmpty: YES / Enveloped: YES / Isolate: SPECIES / Type: VIRUS-LIKE PARTICLE
Buffer solutionpH: 8 / Details: 50mM Tris-HCl pH=8, 60mM NaCl
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMTris-HCl BufferTris-HCl1
260 mMSodium chlorideNaCl1
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Samples of purified HIV-1 RT were diluted to 1mg/mL and 2 molar equivalents of the appropriate compound 12126065 were incubated on a 4oC rotator overnight to combine.
Specimen supportDetails: The cryo-EM grids used for single particle collection were Quantifoil R 2/1 300 gold mesh grids with carbon coating. The grids were glow-discharged for 25 s at 25 mA with the chamber ...Details: The cryo-EM grids used for single particle collection were Quantifoil R 2/1 300 gold mesh grids with carbon coating. The grids were glow-discharged for 25 s at 25 mA with the chamber pressure set at 0.3 mbar (PELCO easiGlow; Ted Pella).
Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 283.15 K
Details: The cryo-EM grids used for single particle collection were Quantifoil R 2/1 300 gold mesh grids with carbon coating. The grids were glow-discharged for 25 s at 25 mA with the chamber ...Details: The cryo-EM grids used for single particle collection were Quantifoil R 2/1 300 gold mesh grids with carbon coating. The grids were glow-discharged for 25 s at 25 mA with the chamber pressure set at 0.3 mbar (PELCO easiGlow; Ted Pella). The grids were prepared using the Leica GP2 plunge freezer with its chamber set to 10oC and 95% humidity. Samples of purified HIV-1 RT were diluted to 1mg/mL and 2 molar equivalents of the appropriate compound 12126065 were incubated on a 4oC rotator overnight to combine. A total of 4uL of HIV-1 RT and compound were applied to the backside of the Quantifoil grid and blotted for 4 seconds on the carbon side before plunge frozen in ethane at -181oC.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.21.1_5286 / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 6268754
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.53 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 175711 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingB value: 125 / Protocol: AB INITIO MODEL / Space: REAL
Atomic model buildingPDB-ID: 4G1Q

4g1q


Accession code: 4G1Q / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 86.21 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00366239
ELECTRON MICROSCOPYf_angle_d0.64028473
ELECTRON MICROSCOPYf_chiral_restr0.044881
ELECTRON MICROSCOPYf_plane_restr0.00571060
ELECTRON MICROSCOPYf_dihedral_angle_d6.7608793

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