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- EMDB-46594: Gag CA-SP1 immature lattice bound with Lenacapavir from enveloped... -

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Basic information

Entry
Database: EMDB / ID: EMD-46594
TitleGag CA-SP1 immature lattice bound with Lenacapavir from enveloped virus like particles
Map dataphysical pixel size 1.068
Sample
  • Virus: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
    • Protein or peptide: Gag polyprotein
  • Ligand: Lenacapavir
  • Ligand: INOSITOL HEXAKISPHOSPHATE
  • Ligand: water
KeywordsHIV-1 / Gag / CA-SP1 / Inhibitor / virion assembly / VIRUS LIKE PARTICLE
Function / homology
Function and homology information


viral budding via host ESCRT complex / ISG15 antiviral mechanism / host multivesicular body / viral nucleocapsid / viral translational frameshifting / host cell nucleus / host cell plasma membrane / virion membrane / structural molecule activity / RNA binding ...viral budding via host ESCRT complex / ISG15 antiviral mechanism / host multivesicular body / viral nucleocapsid / viral translational frameshifting / host cell nucleus / host cell plasma membrane / virion membrane / structural molecule activity / RNA binding / zinc ion binding / membrane
Similarity search - Function
Gag protein p6 / Gag protein p6 / : / gag protein p24 N-terminal domain / Immunodeficiency lentiviral matrix, N-terminal / gag gene protein p17 (matrix protein) / Matrix protein, lentiviral and alpha-retroviral, N-terminal / Retroviral nucleocapsid Gag protein p24, C-terminal domain / Gag protein p24 C-terminal domain / Retrovirus capsid, C-terminal ...Gag protein p6 / Gag protein p6 / : / gag protein p24 N-terminal domain / Immunodeficiency lentiviral matrix, N-terminal / gag gene protein p17 (matrix protein) / Matrix protein, lentiviral and alpha-retroviral, N-terminal / Retroviral nucleocapsid Gag protein p24, C-terminal domain / Gag protein p24 C-terminal domain / 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.
Similarity search - Domain/homology
Biological speciesHuman immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.18 Å
AuthorsWu C / Meuser ME / Xiong Y
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)U54 AI170791 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01 AI178846 United States
CitationJournal: bioRxiv / Year: 2024
Title: Structural insights into inhibitor mechanisms on immature HIV-1 Gag lattice revealed by high-resolution single-particle cryo-EM.
Authors: Chunxiang Wu / Megan E Meuser / Juan S Rey / Hamed Meshkin / Rachel Yang / Swapnil Chandrakant Devarkar / Christian Freniere / Jiong Shi / Christopher Aiken / Juan R Perilla / Yong Xiong /
Abstract: HIV-1 inhibitors, such as Bevirimat (BVM) and Lenacapavir (LEN), block the production and maturation of infectious virions. However, their mechanisms remain unclear due to the absence of high- ...HIV-1 inhibitors, such as Bevirimat (BVM) and Lenacapavir (LEN), block the production and maturation of infectious virions. However, their mechanisms remain unclear due to the absence of high-resolution structures for BVM complexes and LEN's structural data being limited to the mature capsid. Utilizing perforated virus-like particles (VLPs) produced from mammalian cells, we developed an approach to determine cryo-electron microscopy (cryo-EM) structures of HIV-1 with inhibitors. This allowed for the first structural determination of the native immature HIV-1 particle with BVM and LEN bound inside the VLPs at high resolutions. Our findings offer a more accurate model of BVM engaging the Gag lattice and, importantly, demonstrate that LEN not only binds the mature capsid but also targets the immature lattice in a distinct manner. The binding of LEN induces a conformational change in the capsid protein (CA) region and alters the architecture of the Gag lattice, which may affect the maturation process. These insights expand our understanding of the inhibitory mechanisms of BVM and LEN on HIV-1 and provide valuable clues for the design of future inhibitors.
History
DepositionAug 14, 2024-
Header (metadata) releaseOct 30, 2024-
Map releaseOct 30, 2024-
UpdateDec 25, 2024-
Current statusDec 25, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_46594.png

Downloads & links

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Map

FileDownload / File: emd_46594.map.gz / Format: CCP4 / Size: 144.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationphysical pixel size 1.068
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.89 Å/pix.
x 336 pix.
= 300.451 Å		0.89 Å/pix.
x 336 pix.
= 300.451 Å		0.89 Å/pix.
x 336 pix.
= 300.451 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8942 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.035
Minimum - Maximum-0.084357895 - 0.1890539
Average (Standard dev.)0.0030704094 (±0.010748989)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions336336336
Spacing336336336
CellA=B=C: 300.4512 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_46594_msk_1.map
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Additional map: physical pixel size 1.068; Sharpened map

Fileemd_46594_additional_1.map
Annotationphysical pixel size 1.068; Sharpened map
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Half map: physical pixel size 1.068

Fileemd_46594_half_map_1.map
Annotationphysical pixel size 1.068
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Half map: physical pixel size 1.068

Fileemd_46594_half_map_2.map
Annotationphysical pixel size 1.068
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Sample components

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Entire : Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)

EntireName: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
Components
  • Virus: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
    • Protein or peptide: Gag polyprotein
  • Ligand: Lenacapavir
  • Ligand: INOSITOL HEXAKISPHOSPHATE
  • Ligand: water

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Supramolecule #1: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)

SupramoleculeName: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: A codon-optimized Gag plasmid with T8I mutation in the SP1 domain (pCMV-Gag-opt) was expressed in HEK293T cells. The enveloped virion like particles were purified by filtration and ...Details: A codon-optimized Gag plasmid with T8I mutation in the SP1 domain (pCMV-Gag-opt) was expressed in HEK293T cells. The enveloped virion like particles were purified by filtration and ultracentrifugation, and directly used as cryo-EM sample.
NCBI-ID: 11698
Sci species name: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
Sci species strain: Clone pNL4-3 / Virus type: VIRUS-LIKE PARTICLE / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: Yes
Host (natural)Organism: Homo sapiens (human)
Virus shellShell ID: 1 / Name: Gag

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Macromolecule #1: Gag polyprotein

MacromoleculeName: Gag polyprotein / type: protein_or_peptide / ID: 1 / Number of copies: 18 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
Strain: Clone pNL4-3
Molecular weightTheoretical: 25.339037 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: VHQAISPRTL NAWVKVVEEK AFSPEVIPMF SALSEGATPQ DLNTMLNTVG GHQAAMQMLK ETINEEAAEW DRLHPVHAGP IAPGQMREP RGSDIAGTTS TLQEQIGWMT HNPPIPVGEI YKRWIILGLN KIVRMYSPTS ILDIRQGPKE PFRDYVDRFY K TLRAEQAS ...String:
VHQAISPRTL NAWVKVVEEK AFSPEVIPMF SALSEGATPQ DLNTMLNTVG GHQAAMQMLK ETINEEAAEW DRLHPVHAGP IAPGQMREP RGSDIAGTTS TLQEQIGWMT HNPPIPVGEI YKRWIILGLN KIVRMYSPTS ILDIRQGPKE PFRDYVDRFY K TLRAEQAS QEVKNWMTET LLVQNANPDC KTILKALGPG ATLEEMMTAC QGVGGPGHKA RVIAEAMSQV T

UniProtKB: Gag polyprotein

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Macromolecule #2: Lenacapavir

MacromoleculeName: Lenacapavir / type: ligand / ID: 2 / Number of copies: 18 / Formula: QNG
Molecular weightTheoretical: 968.282 Da

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Macromolecule #3: INOSITOL HEXAKISPHOSPHATE

MacromoleculeName: INOSITOL HEXAKISPHOSPHATE / type: ligand / ID: 3 / Number of copies: 7 / Formula: IHP
Molecular weightTheoretical: 660.035 Da
Chemical component information

ChemComp-IHP:
INOSITOL HEXAKISPHOSPHATE

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 294 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
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

Concentration2 mg/mL
BufferpH: 7.4
Component:
ConcentrationNameFormula
10.0 mMTris-hydrocloride
100.0 mMsodium chlorideNaCl
1.0 mMEthylenediaminetetraacetic acid

Details: This is the final buffer in which the enveloped viral like particle was resuspended. The Gag-CA-SP1 lattice is inside the viral like particle and thus not in the direct environment of this buffer.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK III
DetailsA NL-MA/NC DNA was transfected and expressed in Expi293F cells. The enveloped virion like particles were purified by filtration and ultracentrifugation, and directly used as cryo-EM sample.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 81000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:

45975

Final reconstructionApplied symmetry - Point group: C6 (6 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.18 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 1379674
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final 3D classificationNumber classes: 5
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

9cwv


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-9d6d:
Gag CA-SP1 immature lattice bound with Lenacapavir from enveloped virus like particles

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