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- PDB-4usn: The structure of the immature HIV-1 capsid in intact virus partic... -

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Basic information

Entry
Database: PDB / ID: 4usn
TitleThe structure of the immature HIV-1 capsid in intact virus particles at sub-nm resolution
ComponentsP24
KeywordsVIRAL PROTEIN / RETROVIRUS / MATURATION / GAG
Function / homologyRetroviral nucleocapsid Gag protein p24, N-terminal / gag protein p24 N-terminal domain / viral process / viral capsid / Retrovirus capsid, C-terminal / Retrovirus capsid, N-terminal / p24
Function and homology information
Biological speciesHUMAN IMMUNODEFICIENCY VIRUS 1
MethodELECTRON MICROSCOPY / electron tomography / cryo EM / Resolution: 8.8 Å
AuthorsSchur, F.K.M. / Hagen, W.J.H. / Rumlova, M. / Ruml, T. / Mueller, B. / Kraeusslich, H.-G. / Briggs, J.A.G.
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 11, 2014Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 5, 2014Provider: repository / Type: Initial release
Revision 1.1Mar 4, 2015Group: Database references
Revision 1.2Aug 23, 2017Group: Data collection / Category: em_image_scans / em_software
Item: _em_software.fitting_id / _em_software.image_processing_id / _em_software.name

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Structure visualization

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  • Deposited structure unit
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  • Simplified surface model + fitted atomic model
  • EMDB-2706
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Structure viewerMolecule:
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Assembly

Deposited unit
A: P24
B: P24
C: P24
D: P24
E: P24
F: P24
G: P24
H: P24
I: P24
J: P24
K: P24
L: P24
M: P24
N: P24
O: P24
P: P24
Q: P24
R: P24


Theoretical massNumber of molelcules
Total (without water)421,70218
Polymers421,70218
Non-polymers00
Water0
1
A: P24
B: P24
C: P24
D: P24
E: P24
F: P24


Theoretical massNumber of molelcules
Total (without water)140,5676
Polymers140,5676
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
G: P24
H: P24


Theoretical massNumber of molelcules
Total (without water)46,8562
Polymers46,8562
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPQS
3
I: P24
J: P24


Theoretical massNumber of molelcules
Total (without water)46,8562
Polymers46,8562
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPQS
4
K: P24
L: P24


Theoretical massNumber of molelcules
Total (without water)46,8562
Polymers46,8562
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPQS
5
M: P24
N: P24


Theoretical massNumber of molelcules
Total (without water)46,8562
Polymers46,8562
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPQS
6
O: P24
P: P24


Theoretical massNumber of molelcules
Total (without water)46,8562
Polymers46,8562
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPQS
7
Q: P24
R: P24


Theoretical massNumber of molelcules
Total (without water)46,8562
Polymers46,8562
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPQS

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Components

#1: Protein
P24


Mass: 23427.887 Da / Num. of mol.: 18 / Fragment: IMMATURE HIV-1 CAPSID, RESIDUES 4-213 / Source method: isolated from a natural source / Source: (natural) HUMAN IMMUNODEFICIENCY VIRUS 1 / References: UniProt: Q9IVM8
Sequence detailsONLY PARTS OF THE CAPSID (P24) SEQUENCE ARE USED

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: electron tomography

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Sample preparation

ComponentName: INTACT HIV-1 PARTICLES TREATED WITH THE PROTEASE INHIBITOR AMPRENAVIR
Type: VIRUS
Buffer solutionName: 25MM MES PH 6.5, 150MM NACL / pH: 6.5 / Details: 25MM MES PH 6.5, 150MM NACL
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: HOLEY CARBON
VitrificationInstrument: FEI VITROBOT MARK II / Cryogen name: ETHANE
Details: VITRIFICATION 1 -- CRYOGEN- ETHANE, HUMIDITY- 100, INSTRUMENT- FEI VITROBOT MARK III, METHOD- DEGASSED C-FLAT 2- 2-2C GRIDS WERE GLOW DISCHARGED FOR 30 SECONDS AT 20 MA. VIRUS SOLUTION WAS ...Details: VITRIFICATION 1 -- CRYOGEN- ETHANE, HUMIDITY- 100, INSTRUMENT- FEI VITROBOT MARK III, 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 imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS / Date: Aug 13, 2013
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 42000 X / Nominal defocus max: 4000 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm
Specimen holderTilt angle max: 60 ° / Tilt angle min: -45 °
Image recordingElectron dose: 40 e/Å2 / Film or detector model: GATAN MULTISCAN
Radiation wavelengthRelative weight: 1

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Processing

EM software
IDNameCategory
1MDFFmodel fitting
2UCSF Chimeramodel fitting
3AV33D reconstruction
4TOM Toolbox3D reconstruction
CTF correctionDetails: PHASE FLIPPING OF INDIVIDUAL TILTS
SymmetryPoint symmetry: C6 (6 fold cyclic)
3D reconstructionResolution: 8.8 Å / Nominal pixel size: 2.025 Å / Actual pixel size: 2.025 Å
Details: SUBMISSION BASED ON EXPERIMENTAL DATA FROM EMDB EMD -2706. (DEPOSITION ID: 12684).
Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL / Details: METHOD--FLEXIBLE REFINEMENT PROTOCOL--X-RAY,NMR
Atomic model building
IDPDB-ID 3D fitting-ID
11L6N

1l6n

1
23DS2

3ds2

1
RefinementHighest resolution: 8.8 Å
Refinement stepCycle: LAST / Highest resolution: 8.8 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms15120 0 0 0 15120

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