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- EMDB-5455: Cryo-electron tomography of native, trimeric HIV-1 BaL Env in com... -

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Basic information

Entry
Database: EMDB / ID: EMD-5455
TitleCryo-electron tomography of native, trimeric HIV-1 BaL Env in complex with soluble 2-domain CD4
Map dataMolecular structure of native HIV-1 BaL Env trimer in complex with soluble CD4
Sample
  • Sample: Molecular structure of native HIV-1 BaL Env trimer in complex with soluble 2-domain CD4
  • Protein or peptide: Envelope glycoprotein
  • Protein or peptide: soluble 2-domain CD4
KeywordsHIV / Env / sCD4
Biological speciesHuman immunodeficiency virus 1 / Homo sapiens (human)
Methodsubtomogram averaging / cryo EM / Resolution: 24.0 Å
AuthorsTran EEH / Borgnia MJ / Kuybeda O / Schauder DM / Bartesaghi A / Frank GA / Sapiro G / Milne JLS / Subramaniam S
CitationJournal: PLoS Pathog / Year: 2012
Title: Structural mechanism of trimeric HIV-1 envelope glycoprotein activation.
Authors: Erin E H Tran / Mario J Borgnia / Oleg Kuybeda / David M Schauder / Alberto Bartesaghi / Gabriel A Frank / Guillermo Sapiro / Jacqueline L S Milne / Sriram Subramaniam /
Abstract: HIV-1 infection begins with the binding of trimeric viral envelope glycoproteins (Env) to CD4 and a co-receptor on target T-cells. Understanding how these ligands influence the structure of Env is of ...HIV-1 infection begins with the binding of trimeric viral envelope glycoproteins (Env) to CD4 and a co-receptor on target T-cells. Understanding how these ligands influence the structure of Env is of fundamental interest for HIV vaccine development. Using cryo-electron microscopy, we describe the contrasting structural outcomes of trimeric Env binding to soluble CD4, to the broadly neutralizing, CD4-binding site antibodies VRC01, VRC03 and b12, or to the monoclonal antibody 17b, a co-receptor mimic. Binding of trimeric HIV-1 BaL Env to either soluble CD4 or 17b alone, is sufficient to trigger formation of the open quaternary conformation of Env. In contrast, VRC01 locks Env in the closed state, while b12 binding requires a partial opening in the quaternary structure of trimeric Env. Our results show that, despite general similarities in regions of the HIV-1 gp120 polypeptide that contact CD4, VRC01, VRC03 and b12, there are important differences in quaternary structures of the complexes these ligands form on native trimeric Env, and potentially explain differences in the neutralizing breadth and potency of antibodies with similar specificities. From cryo-electron microscopic analysis at ∼9 Å resolution of a cleaved, soluble version of trimeric Env, we show that a structural signature of the open Env conformation is a three-helix motif composed of α-helical segments derived from highly conserved, non-glycosylated N-terminal regions of the gp41 trimer. The three N-terminal gp41 helices in this novel, activated Env conformation are held apart by their interactions with the rest of Env, and are less compactly packed than in the post-fusion, six-helix bundle state. These findings suggest a new structural template for designing immunogens that can elicit antibodies targeting HIV at a vulnerable, pre-entry stage.
History
DepositionAug 6, 2012-
Header (metadata) releaseAug 29, 2012-
Map releaseOct 3, 2012-
UpdateOct 3, 2012-
Current statusOct 3, 2012Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.18
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.18
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
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Supplemental images

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Map

FileDownload / File: emd_5455.map.gz / Format: CCP4 / Size: 3.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMolecular structure of native HIV-1 BaL Env trimer in complex with soluble CD4
Voxel sizeX=Y=Z: 4.1 Å
Density
Contour LevelBy AUTHOR: 0.18 / Movie #1: 0.18
Minimum - Maximum-0.55655253 - 0.81296331
Average (Standard dev.)-0.01021015 (±0.12120042)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions100100100
Spacing100100100
CellA=B=C: 410.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.14.14.1
M x/y/z100100100
origin x/y/z0.0000.0000.000
length x/y/z410.000410.000410.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-150-1500
NX/NY/NZ301301151
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS100100100
D min/max/mean-0.5570.813-0.010

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Supplemental data

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Sample components

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Entire : Molecular structure of native HIV-1 BaL Env trimer in complex wit...

EntireName: Molecular structure of native HIV-1 BaL Env trimer in complex with soluble 2-domain CD4
Components
  • Sample: Molecular structure of native HIV-1 BaL Env trimer in complex with soluble 2-domain CD4
  • Protein or peptide: Envelope glycoprotein
  • Protein or peptide: soluble 2-domain CD4

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Supramolecule #1000: Molecular structure of native HIV-1 BaL Env trimer in complex wit...

SupramoleculeName: Molecular structure of native HIV-1 BaL Env trimer in complex with soluble 2-domain CD4
type: sample / ID: 1000 / Oligomeric state: trimer / Number unique components: 2

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Macromolecule #1: Envelope glycoprotein

MacromoleculeName: Envelope glycoprotein / type: protein_or_peptide / ID: 1 / Name.synonym: Env
Details: Envelope glycoproteins present on the surface of intact virions.
Number of copies: 3 / Oligomeric state: trimer / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Human immunodeficiency virus 1 / Strain: BaL / synonym: HIV-1

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Macromolecule #2: soluble 2-domain CD4

MacromoleculeName: soluble 2-domain CD4 / type: protein_or_peptide / ID: 2 / Name.synonym: sCD4 / Details: soluble cellular CD4 / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Homo sapiens (human) / synonym: human

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging

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Sample preparation

BufferpH: 7.5 / Details: TNE Buffer (10 mM Tris, 150 mM NaCl, 1 mM EDTA)
GridDetails: 200 mesh Quantifoil Multi A
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 77 K / Instrument: FEI VITROBOT MARK III
Method: blot for 6 seconds, at 25 degrees C, 100 percent humidity, blot offset of -2, plunge into an ethane slurry cooled by liquid nitrogen

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Electron microscopy

MicroscopeFEI POLARA 300
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.5 µm / Nominal defocus min: 2.5 µm / Nominal magnification: 34000
Specialist opticsEnergy filter - Name: GATAN GIF / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV
Sample stageSpecimen holder model: GATAN LIQUID NITROGEN / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 °
TemperatureAverage: 81 K
DateOct 25, 2010
Image recordingCategory: CCD / Film or detector model: GENERIC GATAN / Average electron dose: 150 e/Å2
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 24.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: IMOD / Details: Resolution for FSC at 0.5 cut-off is 27 Angstrom
DetailsAverage number of tilts used in the 3D reconstructions: 61. Average tomographic tilt angle increment: 2.

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Atomic model buiding 1

Initial modelPDB ID:

Chain - #0 - Chain ID: C / Chain - #1 - Chain ID: G
SoftwareName: Chimera
DetailsProtocol: Rigid body. Automated fitting procedures
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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