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- EMDB-45975: Gag CA-SP1 immature lattice from intact enveloped virus-like particles -

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Basic information

Entry
Database: EMDB / ID: EMD-45975
TitleGag CA-SP1 immature lattice from intact enveloped virus-like particles
Map datamap, not sharpen; physical pixel size of raw data 1.068
Sample
  • Virus: Human immunodeficiency virus type 1 group M subtype B (isolate NY5) (HIV-1)
    • Protein or peptide: Gag
  • Ligand: INOSITOL HEXAKISPHOSPHATE
  • Ligand: water
KeywordsGag / CA-SP1 / VIRUS LIKE PARTICLE
Function / homology
Function and homology information


Synthesis And Processing Of GAG, GAGPOL Polyproteins / host cellular component / host cell nuclear membrane / Integration of viral DNA into host genomic DNA / Autointegration results in viral DNA circles / Minus-strand DNA synthesis / Plus-strand DNA synthesis / 2-LTR circle formation / Uncoating of the HIV Virion / Vpr-mediated nuclear import of PICs ...Synthesis And Processing Of GAG, GAGPOL Polyproteins / host cellular component / host cell nuclear membrane / Integration of viral DNA into host genomic DNA / Autointegration results in viral DNA circles / Minus-strand DNA synthesis / Plus-strand DNA synthesis / 2-LTR circle formation / Uncoating of the HIV Virion / Vpr-mediated nuclear import of PICs / viral budding via host ESCRT complex / Early Phase of HIV Life Cycle / Integration of provirus / APOBEC3G mediated resistance to HIV-1 infection / Binding and entry of HIV virion / Membrane binding and targetting of GAG proteins / Assembly Of The HIV Virion / Budding and maturation of HIV virion / host multivesicular body / viral nucleocapsid / viral translational frameshifting / 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 group M subtype B (isolate HXB2) / Human immunodeficiency virus type 1 group M subtype B (isolate NY5) (HIV-1)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.42 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
DepositionJul 30, 2024-
Header (metadata) releaseOct 30, 2024-
Map releaseOct 30, 2024-
UpdateOct 30, 2024-
Current statusOct 30, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45975.png

Downloads & links

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Map

FileDownload / File: emd_45975.map.gz / Format: CCP4 / Size: 286.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationmap, not sharpen; physical pixel size of raw data 1.068
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.71 Å/pix.
x 422 pix.
= 300.464 Å		0.71 Å/pix.
x 422 pix.
= 300.464 Å		0.71 Å/pix.
x 422 pix.
= 300.464 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.712 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.015
Minimum - Maximum-0.032819245 - 0.0739607
Average (Standard dev.)0.0016272459 (±0.00498622)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions422422422
Spacing422422422
CellA=B=C: 300.46402 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_45975_msk_1.map
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Half map: physical pixel size of raw data 1.068

Fileemd_45975_half_map_1.map
Annotationphysical pixel size of raw data 1.068
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: physical pixel size of raw data 1.068

Fileemd_45975_half_map_2.map
Annotationphysical pixel size of raw data 1.068
Projections & Slices
AxesZYX

Projections

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Sample components

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Entire : Human immunodeficiency virus type 1 group M subtype B (isolate NY...

EntireName: Human immunodeficiency virus type 1 group M subtype B (isolate NY5) (HIV-1)
Components
  • Virus: Human immunodeficiency virus type 1 group M subtype B (isolate NY5) (HIV-1)
    • Protein or peptide: Gag
  • Ligand: INOSITOL HEXAKISPHOSPHATE
  • Ligand: water

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Supramolecule #1: Human immunodeficiency virus type 1 group M subtype B (isolate NY...

SupramoleculeName: Human immunodeficiency virus type 1 group M subtype B (isolate NY5) (HIV-1)
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 the cryo-EM sample.
NCBI-ID: 11698
Sci species name: Human immunodeficiency virus type 1 group M subtype B (isolate NY5) (HIV-1)
Virus type: VIRUS-LIKE PARTICLE / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: Yes
Host (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Gag

MacromoleculeName: Gag / type: protein_or_peptide / ID: 1 / Number of copies: 18 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus type 1 group M subtype B (isolate HXB2)
Strain: isolate HXB2
Molecular weightTheoretical: 25.500176 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: HQAISPRTLN AWVKVVEEKA FSPEVIPMFS ALSEGATPQD LNTMLNTVGG HQAAMQMLKE TINEEAAEWD RVHPVHAGPI APGQMREPR GSDIAGTTST LQEQIGWMTN NPPIPVGEIY KRWIILGLNK IVRMYSPTSI LDIRQGPKEP FRDYVDRFYK T LRAEQASQ ...String:
HQAISPRTLN AWVKVVEEKA FSPEVIPMFS ALSEGATPQD LNTMLNTVGG HQAAMQMLKE TINEEAAEWD RVHPVHAGPI APGQMREPR GSDIAGTTST LQEQIGWMTN NPPIPVGEIY KRWIILGLNK IVRMYSPTSI LDIRQGPKEP FRDYVDRFYK T LRAEQASQ EVKNWMTETL LVQNANPDCK TILKALGPAA TLEEMMTACQ GVGGPGHKAR VLAEAMSQVI NSA

UniProtKB: Gag polyprotein

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Macromolecule #2: INOSITOL HEXAKISPHOSPHATE

MacromoleculeName: INOSITOL HEXAKISPHOSPHATE / type: ligand / ID: 2 / Number of copies: 7 / Formula: IHP
Molecular weightTheoretical: 660.035 Da
Chemical component information

ChemComp-IHP:
INOSITOL HEXAKISPHOSPHATE

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Macromolecule #3: water

MacromoleculeName: water / type: ligand / ID: 3 / Number of copies: 186 / 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-hydrochloride
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 not directly in this buffer environment.
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Details: 15 mA current
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK III
DetailsIntact enveloped viral-like particles directly purified from human cell culture.

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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 opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.2 µm
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: PDB ENTRY
PDB model - PDB ID:

7asl

Final reconstructionApplied symmetry - Point group: C6 (6 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.42 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Software - details: local refinement / Number images used: 1454696
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC / Software - details: heterogenous refinement
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC / Software - details: local refinement
Final 3D classificationNumber classes: 2
Software:
Namedetails
cryoSPARCheterogenous refinalment
cryoSPARC3D classification
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

7asl


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

PDB-9cwv:
Gag CA-SP1 immature lattice from intact enveloped virus-like particles

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