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- EMDB-2706: The structure of the immature HIV-1 capsid in intact virus partic... -

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Basic information

Entry
Database: EMDB / ID: EMD-2706
TitleThe structure of the immature HIV-1 capsid in intact virus particles at sub-nm resolution
Map dataSubtomogram averaging reconstruction of the immature HIV-1 capsid from intact virus particles
Sample
  • Sample: intact HIV-1 particles treated with the protease inhibitor Amprenavir
  • Virus: Human immunodeficiency virus 1
Keywordsimmature HIV / Retrovirus / Maturation / capsid / Gag
Function / homology: / gag protein p24 N-terminal domain / viral process / Retroviral nucleocapsid Gag protein p24, C-terminal domain / Gag protein p24 C-terminal domain / Retrovirus capsid, C-terminal / Retrovirus capsid, N-terminal / viral capsid / p24
Function and homology information
Biological speciesHuman immunodeficiency virus 1
Methodsubtomogram averaging / cryo EM / Resolution: 8.8 Å
AuthorsSchur FKM / Hagen WJH / Rumlova M / Ruml T / Mueller B / Kraeusslich H-G / Briggs JAG
CitationJournal: Nature / Year: 2015
Title: Structure of the immature HIV-1 capsid in intact virus particles at 8.8 Å resolution.
Authors: Florian K M Schur / Wim J H Hagen / Michaela Rumlová / Tomáš Ruml / Barbara Müller / Hans-Georg Kräusslich / John A G Briggs /
Abstract: Human immunodeficiency virus type 1 (HIV-1) assembly proceeds in two stages. First, the 55 kilodalton viral Gag polyprotein assembles into a hexameric protein lattice at the plasma membrane of the ...Human immunodeficiency virus type 1 (HIV-1) assembly proceeds in two stages. First, the 55 kilodalton viral Gag polyprotein assembles into a hexameric protein lattice at the plasma membrane of the infected cell, inducing budding and release of an immature particle. Second, Gag is cleaved by the viral protease, leading to internal rearrangement of the virus into the mature, infectious form. Immature and mature HIV-1 particles are heterogeneous in size and morphology, preventing high-resolution analysis of their protein arrangement in situ by conventional structural biology methods. Here we apply cryo-electron tomography and sub-tomogram averaging methods to resolve the structure of the capsid lattice within intact immature HIV-1 particles at subnanometre resolution, allowing unambiguous positioning of all α-helices. The resulting model reveals tertiary and quaternary structural interactions that mediate HIV-1 assembly. Strikingly, these interactions differ from those predicted by the current model based on in vitro-assembled arrays of Gag-derived proteins from Mason-Pfizer monkey virus. To validate this difference, we solve the structure of the capsid lattice within intact immature Mason-Pfizer monkey virus particles. Comparison with the immature HIV-1 structure reveals that retroviral capsid proteins, while having conserved tertiary structures, adopt different quaternary arrangements during virus assembly. The approach demonstrated here should be applicable to determine structures of other proteins at subnanometre resolution within heterogeneous environments.
History
DepositionJul 10, 2014-
Header (metadata) releaseAug 6, 2014-
Map releaseNov 5, 2014-
UpdateJan 21, 2015-
Current statusJan 21, 2015Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.27
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.27
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-4usn
  • Surface level: 0.27
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-4usn
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_2706.tif

Downloads & links

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Map

FileDownload / File: emd_2706.map.gz / Format: CCP4 / Size: 10.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram averaging reconstruction of the immature HIV-1 capsid from intact virus particles
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.03 Å/pix.
x 140 pix.
= 283.5 Å		2.03 Å/pix.
x 140 pix.
= 283.5 Å		2.03 Å/pix.
x 140 pix.
= 283.5 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.025 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.27 / Movie #1: 0.27
Minimum - Maximum-0.72606355 - 0.87029272
Average (Standard dev.)-0.00395048 (±0.09804771)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions140140140
Spacing140140140
CellA=B=C: 283.5 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.0252.0252.025
M x/y/z140140140
origin x/y/z0.0000.0000.000
length x/y/z283.500283.500283.500
α/β/γ90.00090.00090.000
start NX/NY/NZ-180-180-179
NX/NY/NZ360360360
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS140140140
D min/max/mean-0.7260.870-0.004

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Supplemental data

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Sample components

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Entire : intact HIV-1 particles treated with the protease inhibitor Amprenavir

EntireName: intact HIV-1 particles treated with the protease inhibitor Amprenavir
Components
  • Sample: intact HIV-1 particles treated with the protease inhibitor Amprenavir
  • Virus: Human immunodeficiency virus 1

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Supramolecule #1000: intact HIV-1 particles treated with the protease inhibitor Amprenavir

SupramoleculeName: intact HIV-1 particles treated with the protease inhibitor Amprenavir
type: sample / ID: 1000 / Oligomeric state: Homohexameric / Number unique components: 1

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Supramolecule #1: Human immunodeficiency virus 1

SupramoleculeName: Human immunodeficiency virus 1 / type: virus / ID: 1 / Name.synonym: HIV-1 / NCBI-ID: 11676 / Sci species name: Human immunodeficiency virus 1 / Virus type: VIRION / Virus isolate: SPECIES / Virus enveloped: Yes / Virus empty: No / Syn species name: HIV-1
Host (natural)Organism: Homo sapiens (human) / synonym: VERTEBRATES

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferpH: 6.5 / Details: 25mM MES pH 6.5, 150mM NaCl
GridDetails: C-Flat 2/2-2C glow discharged
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK II
Method: Degassed C-Flat 2/2-2C grids were glow discharged for 30 seconds at 20 mA. Virus solution was diluted in PBS containing 10nm colloidal gold. 2 ul of this mixture was applied to a grid. Blotting time: 2 seconds

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at nominal working magnification
Specialist opticsEnergy filter - Name: GATAN GIF 2002
DateAug 13, 2013
Image recordingCategory: CCD / Film or detector model: GATAN MULTISCAN / Average electron dose: 40 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 42000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Tilt series - Axis1 - Min angle: -45 ° / Tilt series - Axis1 - Max angle: 60 °
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsSubtomogram averaging calculations were performed using the AV3 and TOM packages. Subtomograms were extracted from the surface of the virus.
Final reconstructionApplied symmetry - Point group: C6 (6 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 8.8 Å / Resolution method: OTHER / Software - Name: TOM, AV3
Details: Reconstruction carried out using subtomogram averaging. Subtomogram averaging was performed using scripts from the TOM (Nickell et al, 2005) and AV3 (Foerster et al, 2005) packages.
Number subtomograms used: 32455
CTF correctionDetails: Phase flipping of individual tilts

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Atomic model buiding 1

Initial modelPDB ID:

1l6n


Chain - Chain ID: 1
SoftwareName: Chimera, MDFF
DetailsStructures for the CA-NTD (PDB 1L6N, chain 1) and CA-CTD (PDB 3DS2, one monomer) were rigid body docked into the EM-density using the "Fit in map" option in chimera. The rigid body fit was further refined using Molecular Dynamics Flexible Fitting.
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-4usn:
The structure of the immature HIV-1 capsid in intact virus particles at sub-nm resolution

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Atomic model buiding 2

Initial modelPDB ID:

3ds2

SoftwareName: Chimera, MDFF
DetailsStructures for the CA-NTD (PDB 1L6N, chain 1) and CA-CTD (PDB 3DS2, one monomer) were rigid body docked into the EM-density using the "Fit in map" option in chimera. The rigid body fit was further refined using Molecular Dynamics Flexible Fitting.
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-4usn:
The structure of the immature HIV-1 capsid in intact virus particles at sub-nm resolution

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