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- EMDB-5447: Molecular Architecture of the Unliganded Membrane-Bound HIV-1 Env... -

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Basic information

Entry
Database: EMDB / ID: EMD-5447
TitleMolecular Architecture of the Unliganded Membrane-Bound HIV-1 Envelope Glycoprotein Trimer
Map dataReconstruction of the unliganded HIV-1 envelope glycoprotein trimer
Sample
  • Sample: HIV-1 envelope glycoprotein trimer
  • Protein or peptide: HIV-1 envelope glycoprotein trimer precursor
KeywordsHIV / envelope spike / trimer / membrane glycoprotein
Biological speciesHuman immunodeficiency virus 1
Methodsingle particle reconstruction / cryo EM / Resolution: 6.0 Å
AuthorsMao Y / Wang L / Sodroski J
CitationJournal: Proc Natl Acad Sci U S A / Year: 2013
Title: Molecular architecture of the uncleaved HIV-1 envelope glycoprotein trimer.
Authors: Youdong Mao / Liping Wang / Christopher Gu / Alon Herschhorn / Anik Désormeaux / Andrés Finzi / Shi-Hua Xiang / Joseph G Sodroski /
Abstract: The human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) trimer, a membrane-fusing machine, mediates virus entry into host cells and is the sole virus-specific target for ...The human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) trimer, a membrane-fusing machine, mediates virus entry into host cells and is the sole virus-specific target for neutralizing antibodies. Binding the receptors, CD4 and CCR5/CXCR4, triggers Env conformational changes from the metastable unliganded state to the fusion-active state. We used cryo-electron microscopy to obtain a 6-Å structure of the membrane-bound, heavily glycosylated HIV-1 Env trimer in its uncleaved and unliganded state. The spatial organization of secondary structure elements reveals that the unliganded conformations of both glycoprotein (gp)120 and gp41 subunits differ from those induced by receptor binding. The gp120 trimer association domains, which contribute to interprotomer contacts in the unliganded Env trimer, undergo rearrangement upon CD4 binding. In the unliganded Env, intersubunit interactions maintain the gp41 ectodomain helical bundles in a "spring-loaded" conformation distinct from the extended helical coils of the fusion-active state. Quaternary structure regulates the virus-neutralizing potency of antibodies targeting the conserved CD4-binding site on gp120. The Env trimer architecture provides mechanistic insights into the metastability of the unliganded state, receptor-induced conformational changes, and quaternary structure-based strategies for immune evasion.
History
DepositionJul 11, 2012-
Header (metadata) releaseAug 8, 2012-
Map releaseJun 12, 2013-
UpdateOct 7, 2015-
Current statusOct 7, 2015Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.007
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.007
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5447_1.tif

Downloads & links

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Map

FileDownload / File: emd_5447.map.gz / Format: CCP4 / Size: 62.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of the unliganded HIV-1 envelope glycoprotein trimer
Voxel sizeX=Y=Z: 0.74688 Å
Density
Contour LevelBy AUTHOR: 0.007 / Movie #1: 0.007
Minimum - Maximum-0.05022446 - 0.07631132
Average (Standard dev.)-0.00001735 (±0.00299701)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 191.20128 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.746878906250.746878906250.74687890625
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z191.201191.201191.201
α/β/γ90.00090.00090.000
start NX/NY/NZ-800-4
NX/NY/NZ1611358
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0500.076-0.000

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Supplemental data

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Sample components

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Entire : HIV-1 envelope glycoprotein trimer

EntireName: HIV-1 envelope glycoprotein trimer
Components
  • Sample: HIV-1 envelope glycoprotein trimer
  • Protein or peptide: HIV-1 envelope glycoprotein trimer precursor

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Supramolecule #1000: HIV-1 envelope glycoprotein trimer

SupramoleculeName: HIV-1 envelope glycoprotein trimer / type: sample / ID: 1000
Details: The sample was monodisperse and solubilized in Cymal-6.
Oligomeric state: three gp120 exterior subunits and three gp41 transmembrane subunits
Number unique components: 3
Molecular weightExperimental: 440 KDa / Theoretical: 435 KDa / Method: Sedimentation, gel electrophoresis

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Macromolecule #1: HIV-1 envelope glycoprotein trimer precursor

MacromoleculeName: HIV-1 envelope glycoprotein trimer precursor / type: protein_or_peptide / ID: 1 / Name.synonym: Env trimer, gp160 trimer
Details: The glycoprotein was expressed in 293F cells and was purified from the extracted plasma membranes. The cDNA sequence is derived from HIV-1 JR-FL isolate.
Oligomeric state: trimer / Recombinant expression: Yes
Source (natural)Organism: Human immunodeficiency virus 1 / synonym: HIV-1
Molecular weightExperimental: 440 KDa / Theoretical: 435 KDa
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant plasmid: pcDNA3.1 in 293F cell line

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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

Concentration2.5 mg/mL
BufferpH: 7.4 / Details: 20 mM Tris-HCl, 300 mM NaCl and 0.01% Cymal-6
GridDetails: 200 mesh and 400 mesh C-flat grid with thin holey carbon support
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 93 K / Instrument: FEI VITROBOT MARK IV
Method: Held for 2 s, blotted by filter papers for 2 s before plunging.

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 200835 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.2 µm / Nominal magnification: 150000
Sample stageSpecimen holder: CT3500 / Specimen holder model: GATAN LIQUID NITROGEN
TemperatureMin: 89 K / Max: 93 K / Average: 91 K
DateSep 1, 2009
Image recordingCategory: CCD / Film or detector model: GENERIC GATAN (4k x 4k) / Number real images: 5991 / Average electron dose: 10 e/Å2
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: Wiener filter
Final angle assignmentDetails: 0.5 degree of angular step
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 6.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER
Details: The final map was deconvoluted and amplitude-corrected with a B-factor of 250 and was low-pass filtered at 5.6 Angstrom with a cosine edge of 8-Fourier-pixel width. The final resolution of ...Details: The final map was deconvoluted and amplitude-corrected with a B-factor of 250 and was low-pass filtered at 5.6 Angstrom with a cosine edge of 8-Fourier-pixel width. The final resolution of the refined cryo-EM map, measured by FSC-0.5 cutoff, is 6 Angstrom without masking of the background noise in the map and is 5.66 Angstrom with of masking the background noise.
Number images used: 582914

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Atomic model buiding 1

Initial modelPDB ID:

3jwd


Chain - Chain ID: G
SoftwareName: O, Coot, CNS, Chimera, Modeller
DetailsProtocol: Flexible fitting
RefinementSpace: REAL / Protocol: FLEXIBLE FIT

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