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- EMDB-45763: HIV-2 CA pentamer in the presence of CPSF6 peptide but without bi... -

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Basic information

Entry
Database: EMDB / ID: EMD-45763
TitleHIV-2 CA pentamer in the presence of CPSF6 peptide but without binding; assembled via liposome templating
Map dataEM map of HIV-2 CA pentamer assembled via templating on functionalized liposomes. CPSF6 peptide was introduced, but no density is observed.
Sample
  • Complex: HIV-2 capsid protein assembled into a lattice via liposome templating and then bound with CPSF6 peptide.
    • Complex: HIV-2 capsid protein
    • Complex: CPSF6 peptide
    • Protein or peptide: HIV-2 capsid protein
    • Protein or peptide: CPSF6
KeywordsHIV-2 / Capsid / IP6 / CPSF6 / VIRAL PROTEIN
Biological speciesHuman immunodeficiency virus 2 / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.82 Å
AuthorsFreniere C / Cook M / Xiong Y
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)T32GM008283 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)U54AI170791 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)P50AI150481 United States
CitationJournal: Cell Rep / Year: 2025
Title: Structural insights into HIV-2 CA lattice formation and FG-pocket binding revealed by single-particle cryo-EM.
Authors: Matthew Cook / Christian Freniere / Chunxiang Wu / Faith Lozano / Yong Xiong /
Abstract: One of the striking features of human immunodeficiency virus (HIV) is the capsid, a fullerene cone comprised of pleomorphic capsid protein (CA) that shields the viral genome and recruits cofactors. ...One of the striking features of human immunodeficiency virus (HIV) is the capsid, a fullerene cone comprised of pleomorphic capsid protein (CA) that shields the viral genome and recruits cofactors. Despite significant advances in understanding the mechanisms of HIV-1 CA assembly and host factor interactions, HIV-2 CA assembly remains poorly understood. By templating the assembly of HIV-2 CA on functionalized liposomes, we report high-resolution structures of the HIV-2 CA lattice, including both CA hexamers and pentamers, alone and with peptides of host phenylalanine-glycine (FG)-motif proteins Nup153 and CPSF6. While the overall fold and mode of FG-peptide binding is conserved with HIV-1, this study reveals distinctive features of the HIV-2 CA lattice, including differing structural character at regions of host factor interactions and divergence in the mechanism of formation of CA hexamers and pentamers. This study extends our understanding of HIV capsids and highlights an approach facilitating the study of lentiviral capsid biology.
History
DepositionJul 15, 2024-
Header (metadata) releaseMar 5, 2025-
Map releaseMar 5, 2025-
UpdateMar 5, 2025-
Current statusMar 5, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45763.png

Downloads & links

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Map

FileDownload / File: emd_45763.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationEM map of HIV-2 CA pentamer assembled via templating on functionalized liposomes. CPSF6 peptide was introduced, but no density is observed.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 180 pix.
= 192.24 Å		1.07 Å/pix.
x 180 pix.
= 192.24 Å		1.07 Å/pix.
x 180 pix.
= 192.24 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.068 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-0.17854796 - 0.538127
Average (Standard dev.)0.011789161 (±0.038060207)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 192.23999 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: EM half map B of HIV-2 CA pentamer...

Fileemd_45763_half_map_1.map
AnnotationEM half map B of HIV-2 CA pentamer in the presence of CPSF6 peptide, though no density for the peptide is observed.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: EM half map A of HIV-2 CA pentamer...

Fileemd_45763_half_map_2.map
AnnotationEM half map A of HIV-2 CA pentamer in the presence of CPSF6 peptide, though no density for the peptide is observed.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : HIV-2 capsid protein assembled into a lattice via liposome templa...

EntireName: HIV-2 capsid protein assembled into a lattice via liposome templating and then bound with CPSF6 peptide.
Components
  • Complex: HIV-2 capsid protein assembled into a lattice via liposome templating and then bound with CPSF6 peptide.
    • Complex: HIV-2 capsid protein
    • Complex: CPSF6 peptide
    • Protein or peptide: HIV-2 capsid protein
    • Protein or peptide: CPSF6

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Supramolecule #1: HIV-2 capsid protein assembled into a lattice via liposome templa...

SupramoleculeName: HIV-2 capsid protein assembled into a lattice via liposome templating and then bound with CPSF6 peptide.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: C-terminally hexahistidine tagged HIV-2 CA associated with a liposome decorated with NiNTA headgroups which results in the assembly of a lattice of CA. CPSF6 peptide was then introduced and ...Details: C-terminally hexahistidine tagged HIV-2 CA associated with a liposome decorated with NiNTA headgroups which results in the assembly of a lattice of CA. CPSF6 peptide was then introduced and allowed to equilibrate binding.
Molecular weightTheoretical: 26.9 KDa

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Supramolecule #2: HIV-2 capsid protein

SupramoleculeName: HIV-2 capsid protein / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: Human immunodeficiency virus 2 / Strain: GL-AN

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Supramolecule #3: CPSF6 peptide

SupramoleculeName: CPSF6 peptide / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Homo sapiens (human) / Synthetically produced: Yes

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Macromolecule #1: HIV-2 capsid protein

MacromoleculeName: HIV-2 capsid protein / type: protein_or_peptide / ID: 1
Details: HIV-2 capsid protein with C-terminal Gly-Ser-Ser linker and hexahistidine tag following proteolytic processing of the N-terminal Met.
Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 2 / Strain: GL-AN
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: PVQQTGGGNY IHVPLSPRTL NAWVKLVEDK KFGAEVVPGF QALSEGCTPY DINQMLNCVG DHQAAMQIIR EIINDEAADW DAQHPIPGPL PAGQLRDPRG SDIAGTTSTV EEQIQWMYRP QNPVPVGNIY RRWIQIGLQK CVRMYNPTNI LDVKQGPKEP FQSYVDRFYK ...String:
PVQQTGGGNY IHVPLSPRTL NAWVKLVEDK KFGAEVVPGF QALSEGCTPY DINQMLNCVG DHQAAMQIIR EIINDEAADW DAQHPIPGPL PAGQLRDPRG SDIAGTTSTV EEQIQWMYRP QNPVPVGNIY RRWIQIGLQK CVRMYNPTNI LDVKQGPKEP FQSYVDRFYK SLRAEQTDPA VKNWMTQTLL IQNANPDCKL VLKGLGMNPT LEEMLTACQG VGGPGQKARL MGSSHHHHHH

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Macromolecule #2: CPSF6

MacromoleculeName: CPSF6 / type: protein_or_peptide / ID: 2 / Details: Truncation peptide of CPSF6 (313-327) / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
SequenceString:
PVLFPGQPFG QPPLG

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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

Concentration10.7 mg/mL
BufferpH: 7
Component:
ConcentrationNameFormula
20.0 mMHEPES
250.0 mMSodium chlorideNaCl
0.1 mMTCEP
4.0 mMIP6

Details: The mixed buffer of storage buffer for the protein and lipid components with IP6 supplemented.
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Support film - Film thickness: 12 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.015 kPa / Details: 15 mA discharge current.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV
Details: Grids were dual-side blotted with blot force 0 for 5.5 sec before plunge freezing in liquid ethane..
DetailsSample was prepared with 400 uM HIV-2 CA-6xHis protein, 5.9 mM lipid mix (described in publication), and 4 mM IP6 as final concentrations following subsequent addition of CPSF6 peptide. After assembly, CPSF6 was added to a final concentration of 400 uM. Sample was well-distributed on the grid, mostly monodisperse. Perhaps slightly more particles on carbon versus in the hole.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 15 eV
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 opticsC2 aperture diameter: 30.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 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

Particle selectionNumber selected: 23033474
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C5 (5 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.82 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 1906465
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationSoftware - Name: cryoSPARC
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

9clj


Chain - Chain ID: A / Chain - Residue range: 1-223 / Chain - Source name: Other / Chain - Initial model type: experimental model
Details: NTDs matched well, but the CTD had to be realigned.
DetailsThe HIV-2 CA pentamer chain derived from micelle-templated icosahedra described in this publication was used for rigid fitting into the density for confidence in the similarity between peptide-less datasets.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation coefficient

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