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- EMDB-49521: HIV-1 Reverse Transcriptase with New Non-Nucleoside Reverse Trans... -

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Basic information

Entry
Database: EMDB / ID: EMD-49521
TitleHIV-1 Reverse Transcriptase with New Non-Nucleoside Reverse Transcriptase Inhibitor 12126065
Map data
Sample
  • Organelle or cellular component: HIV-1 reverse transcriptase with novel inhibitor
    • Protein or peptide: Reverse transcriptase p66 subunit
    • Protein or peptide: Reverse transcriptase p51 subunit
  • Ligand: 4-({5-amino-1-[6-(2-cyanoethyl)naphthalene-1-sulfonyl]-1H-1,2,4-triazol-3-yl}amino)-2-chlorobenzonitrile
KeywordsReverse transcriptase / VIRAL PROTEIN
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
Biological speciesHIV-1 06TG.HT008 (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.53 Å
AuthorsYoung MA / Lane TR / Raman R / Nelson JAE / Riabova O / Kazakova E / Monakhova N / Tsedilin A / Rees SD / Quinnell D ...Young MA / Lane TR / Raman R / Nelson JAE / Riabova O / Kazakova E / Monakhova N / Tsedilin A / Rees SD / Quinnell D / Chang G / Ekins S
Funding support1 items
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, 2025-
Header (metadata) releaseMay 7, 2025-
Map releaseMay 7, 2025-
UpdateMay 21, 2025-
Current statusMay 21, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_49521.png

Downloads & links

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Map

FileDownload / File: emd_49521.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 256 pix.
= 215.04 Å		0.84 Å/pix.
x 256 pix.
= 215.04 Å		0.84 Å/pix.
x 256 pix.
= 215.04 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.84 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.042
Minimum - Maximum-0.5258282 - 0.88315666
Average (Standard dev.)0.00002730252 (±0.018441383)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 215.04 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_49521_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_49521_half_map_2.map
Projections & Slices
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Histogram

Histogram (log scale)

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Sample components

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Entire : HIV-1 reverse transcriptase with novel inhibitor

EntireName: HIV-1 reverse transcriptase with novel inhibitor
Components
  • Organelle or cellular component: HIV-1 reverse transcriptase with novel inhibitor
    • Protein or peptide: Reverse transcriptase p66 subunit
    • Protein or peptide: Reverse transcriptase p51 subunit
  • Ligand: 4-({5-amino-1-[6-(2-cyanoethyl)naphthalene-1-sulfonyl]-1H-1,2,4-triazol-3-yl}amino)-2-chlorobenzonitrile

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Supramolecule #1: HIV-1 reverse transcriptase with novel inhibitor

SupramoleculeName: HIV-1 reverse transcriptase with novel inhibitor / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: HIV-1 06TG.HT008 (virus)
Molecular weightTheoretical: 1.17 MDa

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Macromolecule #1: Reverse transcriptase p66 subunit

MacromoleculeName: Reverse transcriptase p66 subunit / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: RNA-directed DNA polymerase
Source (natural)Organism: HIV-1 06TG.HT008 (virus)
Molecular weightTheoretical: 65.8845 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MPISPIETVP VKLKPGMDGP KVKQWPLTEE KIKALVEICT EMEKEGKISK IGPENPYNTP VFAIKKKDST KWRKLVDFRE LNKRTQDFW EVQLGIPHPA GLKQKKSVTV LDVGDAYFSV PLDKDFRKYT AFTIPSINNE TPGIRYQYNV LPQGWKGSPA I FQCSMTKI ...String:
MPISPIETVP VKLKPGMDGP KVKQWPLTEE KIKALVEICT EMEKEGKISK IGPENPYNTP VFAIKKKDST KWRKLVDFRE LNKRTQDFW EVQLGIPHPA GLKQKKSVTV LDVGDAYFSV PLDKDFRKYT AFTIPSINNE TPGIRYQYNV LPQGWKGSPA I FQCSMTKI LEPFRKQNPD IVIYQYMDDL YVGSDLEIGQ HRTKIEELRQ HLLRWGFTTP DKKHQKEPPF LWMGYELHPD KW TVQPIVL PEKDSWTVND IQKLVGKLNW ASQIYAGIKV RQLCKLLRGT KALTEVVPLT EEAELELAEN REILKEPVHG VYY DPSKDL IAEIQKQGQG QWTYQIYQEP FKNLKTGKYA RMKGAHTNDV KQLTEAVQKI ATESIVIWGK TPKFKLPIQK ETWE AWWTE YWQATWIPEW EFVNTPPLVK LWYQLEKEPI IGAETFYVDG AANRETKLGK AGYVTDRGRQ KVVPLTDTTN QKTEL QAIH LALQDSGLEV NIVTDSQYAL GIIQAQPDKS ESELVSQIIE QLIKKEKVYL AWVPAHKGIG GNEQVDGLVS AGIRKV LHH HHHHHHHH

UniProtKB: Gag-Pol polyprotein

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Macromolecule #2: Reverse transcriptase p51 subunit

MacromoleculeName: Reverse transcriptase p51 subunit / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: RNA-directed DNA polymerase
Source (natural)Organism: HIV-1 06TG.HT008 (virus)
Molecular weightTheoretical: 51.465234 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MPISPIETVP VKLKPGMDGP KVKQWPLTEE KIKALVEICT EMEKEGKISK IGPENPYNTP VFAIKKKDST KWRKLVDFRE LNKRTQDFW EVQLGIPHPA GLKQKKSVTV LDVGDAYFSV PLDKDFRKYT AFTIPSINNE TPGIRYQYNV LPQGWKGSPA I FQCSMTKI ...String:
MPISPIETVP VKLKPGMDGP KVKQWPLTEE KIKALVEICT EMEKEGKISK IGPENPYNTP VFAIKKKDST KWRKLVDFRE LNKRTQDFW EVQLGIPHPA GLKQKKSVTV LDVGDAYFSV PLDKDFRKYT AFTIPSINNE TPGIRYQYNV LPQGWKGSPA I FQCSMTKI LEPFRKQNPD IVIYQYMDDL YVGSDLEIGQ HRTKIEELRQ HLLRWGFTTP DKKHQKEPPF LWMGYELHPD KW TVQPIVL PEKDSWTVND IQKLVGKLNW ASQIYAGIKV RQLCKLLRGT KALTEVVPLT EEAELELAEN REILKEPVHG VYY DPSKDL IAEIQKQGQG QWTYQIYQEP FKNLKTGKYA RMKGAHTNDV KQLTEAVQKI ATESIVIWGK TPKFKLPIQK ETWE AWWTE YWQATWIPEW EFVNTPPLVK LWYQLEKEPI IGAETF

UniProtKB: Gag-Pol polyprotein

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Macromolecule #3: 4-({5-amino-1-[6-(2-cyanoethyl)naphthalene-1-sulfonyl]-1H-1,2,4-t...

MacromoleculeName: 4-({5-amino-1-[6-(2-cyanoethyl)naphthalene-1-sulfonyl]-1H-1,2,4-triazol-3-yl}amino)-2-chlorobenzonitrile
type: ligand / ID: 3 / Number of copies: 1 / Formula: A1BYY
Molecular weightTheoretical: 477.926 Da

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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

Concentration1 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
50.0 mMTris-HClTris-HCl Buffer
60.0 mMNaClSodium chloride

Details: 50mM Tris-HCl pH=8, 60mM NaCl
GridModel: Quantifoil R2/1 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 25 sec. / Pretreatment - Atmosphere: OTHER / Pretreatment - Pressure: 30.0 kPa
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).
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 283.15 K / Instrument: LEICA EM GP
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..
DetailsSamples 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.

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 6268754
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

4g1q

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.53 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 175711
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final 3D classificationNumber classes: 1

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Atomic model buiding 1

Initial modelPDB ID:

4g1q


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 125
Output model

PDB-9nlp:
HIV-1 Reverse Transcriptase with New Non-Nucleoside Reverse Transcriptase Inhibitor 12126065

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