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- EMDB-4422: Representative tomogram of immature MLV particles -

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Basic information

Entry
Database: EMDB / ID: EMD-4422
TitleRepresentative tomogram of immature MLV particles
Map dataRepresentative tomogram of immature MLV particles for subtomogram averaging
Sample
  • Virus: Murine leukemia virus
Biological speciesMurine leukemia virus
Methodelectron tomography / cryo EM
AuthorsQu K / Glass B / Dolezal M / Schur FKM / Rein A / Rumlova M / Ruml T / Kraeusslich HG / Briggs JAG
CitationJournal: Proc Natl Acad Sci U S A / Year: 2018
Title: Structure and architecture of immature and mature murine leukemia virus capsids.
Authors: Kun Qu / Bärbel Glass / Michal Doležal / Florian K M Schur / Brice Murciano / Alan Rein / Michaela Rumlová / Tomáš Ruml / Hans-Georg Kräusslich / John A G Briggs /
Abstract: Retroviruses assemble and bud from infected cells in an immature form and require proteolytic maturation for infectivity. The CA (capsid) domains of the Gag polyproteins assemble a protein lattice as ...Retroviruses assemble and bud from infected cells in an immature form and require proteolytic maturation for infectivity. The CA (capsid) domains of the Gag polyproteins assemble a protein lattice as a truncated sphere in the immature virion. Proteolytic cleavage of Gag induces dramatic structural rearrangements; a subset of cleaved CA subsequently assembles into the mature core, whose architecture varies among retroviruses. Murine leukemia virus (MLV) is the prototypical γ-retrovirus and serves as the basis of retroviral vectors, but the structure of the MLV CA layer is unknown. Here we have combined X-ray crystallography with cryoelectron tomography to determine the structures of immature and mature MLV CA layers within authentic viral particles. This reveals the structural changes associated with maturation, and, by comparison with HIV-1, uncovers conserved and variable features. In contrast to HIV-1, most MLV CA is used for assembly of the mature core, which adopts variable, multilayered morphologies and does not form a closed structure. Unlike in HIV-1, there is similarity between protein-protein interfaces in the immature MLV CA layer and those in the mature CA layer, and structural maturation of MLV could be achieved through domain rotations that largely maintain hexameric interactions. Nevertheless, the dramatic architectural change on maturation indicates that extensive disassembly and reassembly are required for mature core growth. The core morphology suggests that wrapping of the genome in CA sheets may be sufficient to protect the MLV ribonucleoprotein during cell entry.
History
DepositionNov 14, 2018-
Header (metadata) releaseDec 5, 2018-
Map releaseDec 5, 2018-
UpdateDec 19, 2018-
Current statusDec 19, 2018Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
  • Download
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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Movie viewer
Supplemental images

emd_4422.png

Downloads & links

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Map

FileDownload / File: emd_4422.map.gz / Format: CCP4 / Size: 5.2 GB / Type: IMAGE STORED AS SIGNED INTEGER (2 BYTES)
AnnotationRepresentative tomogram of immature MLV particles for subtomogram averaging
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.7 Å/pix.
x 810 pix.
= 2187. Å		2.7 Å/pix.
x 1855 pix.
= 5008.5 Å		2.7 Å/pix.
x 1855 pix.
= 5008.5 Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 2.7 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-1127. - 1224.
Average (Standard dev.)6.217764 (±186.0428)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-1-1-1
Dimensions18551855810
Spacing18551855810
CellA: 5008.5 Å / B: 5008.5 Å / C: 2187.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Integer*27
Å/pix. X/Y/Z2.72.72.7
M x/y/z18551855810
origin x/y/z0.0000.0000.000
length x/y/z5008.5005008.5002187.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-1-1-1
NC/NR/NS18551855810
D min/max/mean-1127.0001224.0006.218

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Supplemental data

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Sample components

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Entire : Murine leukemia virus

EntireName: Murine leukemia virus
Components
  • Virus: Murine leukemia virus

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Supramolecule #1: Murine leukemia virus

SupramoleculeName: Murine leukemia virus / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1 / NCBI-ID: 11786 / Sci species name: Murine leukemia virus / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: No
Host systemOrganism: Homo sapiens (human) / Recombinant cell: HEK 293T / Recombinant plasmid: M2204

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation stateparticle

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Sample preparation

BufferpH: 6
GridModel: C-flat-2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 288 K / Instrument: FEI VITROBOT MARK II
DetailsPurified virus solution was inactivated and diluted 1:1 with PBS containing 10 nm colloidal gold.
SectioningOther: NO SECTIONING
Fiducial markerManufacturer: UMC Utrecht / Diameter: 10 nm

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Digitization - Sampling interval: 5.0 µm / Digitization - Frames/image: 1-6 / Number grids imaged: 1 / Average exposure time: 0.6 sec. / Average electron dose: 2.5 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: 4.5 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 105000
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

Final reconstructionSoftware - Name: eTomo / Number images used: 40
CTF correctionSoftware:
Namedetails
MATLABCTF determination
IMODPhase flipping only

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