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- PDB-9d6c: Gag CA-SP1 immature lattice bound with Lenacapavir and Bevirimat ... -

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Basic information

Entry
Database: PDB / ID: 9d6c
TitleGag CA-SP1 immature lattice bound with Lenacapavir and Bevirimat from enveloped virus like particles
ComponentsGag
KeywordsVIRUS LIKE PARTICLE / HIV-1 / Gag / CA-SP1 / Inhibitor / virion assembly
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
Chem-2I4 / INOSITOL HEXAKISPHOSPHATE / Chem-QNG / Gag polyprotein
Similarity search - Component
Biological speciesHuman immunodeficiency virus type 1 group M subtype B
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.1 Å
AuthorsWu, C. / Meuser, M.E. / Xiong, Y.
Funding support United States, 2items
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, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 30, 2024Provider: repository / Type: Initial release
Revision 1.1Dec 25, 2024Group: Data collection / Derived calculations / Structure summary
Category: chem_comp / em_admin ...chem_comp / em_admin / entity / pdbx_entity_nonpoly
Item: _chem_comp.name / _chem_comp.pdbx_synonyms ..._chem_comp.name / _chem_comp.pdbx_synonyms / _em_admin.last_update / _entity.pdbx_description / _pdbx_entity_nonpoly.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Gag
B: Gag
C: Gag
D: Gag
E: Gag
F: Gag
G: Gag
H: Gag
I: Gag
J: Gag
K: Gag
L: Gag
M: Gag
N: Gag
O: Gag
P: Gag
Q: Gag
R: Gag
hetero molecules


Theoretical massNumber of molelcules
Total (without water)488,75250
Polymers462,60918
Non-polymers26,14332
Water6,810378
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
Gag


Mass: 25700.475 Da / Num. of mol.: 18 / Fragment: CA-SP1 domains (UNP residues 141-372) / Mutation: T239I
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human immunodeficiency virus type 1 group M subtype B (isolate HXB2)
Strain: isolate HXB2 / Gene: gag / Plasmid: pCMV-Gag-opt / Cell line (production host): HEK293T / Production host: Homo sapiens (human) / References: UniProt: P04591
#2: Chemical
ChemComp-QNG / Lenacapavir / Sunlenca / GS-CA1 / GS-6207


Mass: 968.282 Da / Num. of mol.: 18 / Source method: obtained synthetically / Formula: C39H32ClF10N7O5S2 / Feature type: SUBJECT OF INVESTIGATION / Comment: medication, antiretroviral*YM
#3: Chemical
ChemComp-2I4 / 3alpha-[(3-carboxy-3-methylbutanoyl)oxy]-8alpha,9beta,10alpha,13alpha,17alpha,19beta-lup-20(29)-en-28-oic acid / Bevirimat


Mass: 584.826 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: C36H56O6 / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical
ChemComp-IHP / INOSITOL HEXAKISPHOSPHATE / MYO-INOSITOL HEXAKISPHOSPHATE / INOSITOL 1,2,3,4,5,6-HEXAKISPHOSPHATE


Mass: 660.035 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Formula: C6H18O24P6 / Feature type: SUBJECT OF INVESTIGATION
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 378 / Source method: isolated from a natural source / Formula: H2O
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: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
Type: VIRUS
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.
Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)
Strain: Clone pNL4-3
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293T / Plasmid: pCMV-Gag-opt
Details of virusEmpty: YES / Enveloped: YES / Isolate: STRAIN / Type: VIRUS-LIKE PARTICLE
Natural hostOrganism: Homo sapiens
Virus shellName: Gag
Buffer solutionpH: 7.4
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.
Buffer component
IDConc.NameFormulaBuffer-ID
110 mMTris-hydrocloride1
2100 mMsodium chlorideNaCl1
31 mMEthylenediaminetetraacetic acid1
SpecimenConc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
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.
Specimen supportDetails: 15 mA current / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K

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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 FIELD / Nominal magnification: 81000 X / Nominal defocus max: 2200 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM software
IDNameVersionCategoryDetails
2SerialEMimage acquisition
4cryoSPARCCTF correctionpatch CTF estimation
5cryoSPARCCTF correctionglobal CTF refinement
6cryoSPARCCTF correctionlocal CTF refinement
9UCSF ChimeraXmodel fitting
11cryoSPARCinitial Euler assignmentheterogenous refinement
12cryoSPARCfinal Euler assignmentlocal refinement
13cryoSPARCclassificationheterogenous refinement
14cryoSPARCclassification3D classification
15cryoSPARC3D reconstructionlocal refinement
16cryoSPARC3D reconstructionhomogenous reconstruction only
17PHENIX1.21_5207model refinement
18Cootmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C6 (6 fold cyclic)
3D reconstructionResolution: 2.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 1323040 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingPDB-ID: 9CWV

9cwv


Accession code: 9CWV / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 68.45 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.004235110
ELECTRON MICROSCOPYf_angle_d0.708348276
ELECTRON MICROSCOPYf_chiral_restr0.04345218
ELECTRON MICROSCOPYf_plane_restr0.00466064
ELECTRON MICROSCOPYf_dihedral_angle_d13.71455049

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