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- EMDB-8730: Cryo-EM reconstruction of the HIV-1 BG505 SOSIP.664 Env trimer in... -

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Basic information

Entry
Database: EMDB / ID: EMD-8730
TitleCryo-EM reconstruction of the HIV-1 BG505 SOSIP.664 Env trimer in complex with soluble CD4 (D1-D2) and fragment antigen binding variable domain of 17b
Map dataComplex formed by HIV-1 BG505 SOSIP.664 Env with soluble CD4 (2-domain) and broadly neutralizing antibody 17b
Sample
  • Complex: HIV-1 Env BG505 SOSIP.664 in complex with soluble CD4 (2-domain) and Fab domain of neutralizing 17b antibody
    • Complex: HIV-1 Env BG505 SOSIP.664
      • Protein or peptide: HIV-1 Env BG505 SOSIP.664 gp41
      • Protein or peptide: HIV-1 Env BG505 SOSIP.664 gp120
    • Complex: CD4 (2-domain)
      • Protein or peptide: soluble CD4 (2-domain)
    • Complex: broadly neutralizing antibody (Fab) 17b
      • Protein or peptide: broadly neutralizing antibody (Fab) 17b light chain
      • Protein or peptide: broadly neutralizing antibody (Fab) 17b heavy chain
Function / homology
Function and homology information


positive regulation of plasma membrane raft polarization / positive regulation of receptor clustering / positive regulation of establishment of T cell polarity / virus-mediated perturbation of host defense response / host cell endosome membrane / clathrin-dependent endocytosis of virus by host cell / viral protein processing / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope ...positive regulation of plasma membrane raft polarization / positive regulation of receptor clustering / positive regulation of establishment of T cell polarity / virus-mediated perturbation of host defense response / host cell endosome membrane / clathrin-dependent endocytosis of virus by host cell / viral protein processing / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / structural molecule activity / virion attachment to host cell / host cell plasma membrane / virion membrane / plasma membrane
Similarity search - Function
Envelope glycoprotein Gp160 / Retroviral envelope protein / Retroviral envelope protein GP41-like / Gp120 core superfamily / Envelope glycoprotein GP120 / Human immunodeficiency virus 1, envelope glycoprotein Gp120
Similarity search - Domain/homology
Envelope glycoprotein gp160 / Envelope glycoprotein gp160
Similarity search - Component
Biological speciesHuman immunodeficiency virus 1 / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 8.6 Å
AuthorsLyumkis D / de Val N / Ward AB
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)UM1 AI100663 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P50 GM103368 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)P01 AI110657 United States
CitationJournal: Nature / Year: 2017
Title: Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.
Authors: Gabriel Ozorowski / Jesper Pallesen / Natalia de Val / Dmitry Lyumkis / Christopher A Cottrell / Jonathan L Torres / Jeffrey Copps / Robyn L Stanfield / Albert Cupo / Pavel Pugach / John P ...Authors: Gabriel Ozorowski / Jesper Pallesen / Natalia de Val / Dmitry Lyumkis / Christopher A Cottrell / Jonathan L Torres / Jeffrey Copps / Robyn L Stanfield / Albert Cupo / Pavel Pugach / John P Moore / Ian A Wilson / Andrew B Ward /
Abstract: For many enveloped viruses, binding to a receptor(s) on a host cell acts as the first step in a series of events culminating in fusion with the host cell membrane and transfer of genetic material for ...For many enveloped viruses, binding to a receptor(s) on a host cell acts as the first step in a series of events culminating in fusion with the host cell membrane and transfer of genetic material for replication. The envelope glycoprotein (Env) trimer on the surface of HIV is responsible for receptor binding and fusion. Although Env can tolerate a high degree of mutation in five variable regions (V1-V5), and also at N-linked glycosylation sites that contribute roughly half the mass of Env, the functional sites for recognition of receptor CD4 and co-receptor CXCR4/CCR5 are conserved and essential for viral fitness. Soluble SOSIP Env trimers are structural and antigenic mimics of the pre-fusion native, surface-presented Env, and are targets of broadly neutralizing antibodies. Thus, they are attractive immunogens for vaccine development. Here we present high-resolution cryo-electron microscopy structures of subtype B B41 SOSIP Env trimers in complex with CD4 and antibody 17b, or with antibody b12, at resolutions of 3.7 Å and 3.6 Å, respectively. We compare these to cryo-electron microscopy reconstructions of B41 SOSIP Env trimers with no ligand or in complex with either CD4 or the CD4-binding-site antibody PGV04 at 5.6 Å, 5.2 Å and 7.4 Å resolution, respectively. Consequently, we present the most complete description yet, to our knowledge, of the CD4-17b-induced intermediate and provide the molecular basis of the receptor-binding-induced conformational change required for HIV-1 entry into host cells. Both CD4 and b12 induce large, previously uncharacterized conformational rearrangements in the gp41 subunits, and the fusion peptide becomes buried in a newly formed pocket. These structures provide key details on the biological function of the type I viral fusion machine from HIV-1 as well as new templates for inhibitor design.
History
Header (metadata) releaseJan 11, 2017-
DepositionMay 13, 2017-
Map releaseJul 26, 2017-
UpdateJan 29, 2020-
Current statusJan 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.12
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.12
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8730.png

Downloads & links

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Map

FileDownload / File: emd_8730.map.gz / Format: CCP4 / Size: 8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationComplex formed by HIV-1 BG505 SOSIP.664 Env with soluble CD4 (2-domain) and broadly neutralizing antibody 17b
Voxel sizeX=Y=Z: 2.62 Å
Density
Contour LevelBy AUTHOR: 0.12 / Movie #1: 0.12
Minimum - Maximum-0.09550767 - 0.30681568
Average (Standard dev.)-0.0008014025 (±0.018791849)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions128128128
Spacing128128128
CellA=B=C: 335.36 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.622.622.62
M x/y/z128128128
origin x/y/z0.0000.0000.000
length x/y/z335.360335.360335.360
α/β/γ90.00090.00090.000
start NX/NY/NZ-38-19-20
NX/NY/NZ858082
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS128128128
D min/max/mean-0.0960.307-0.001

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Supplemental data

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Sample components

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Entire : HIV-1 Env BG505 SOSIP.664 in complex with soluble CD4 (2-domain) ...

EntireName: HIV-1 Env BG505 SOSIP.664 in complex with soluble CD4 (2-domain) and Fab domain of neutralizing 17b antibody
Components
  • Complex: HIV-1 Env BG505 SOSIP.664 in complex with soluble CD4 (2-domain) and Fab domain of neutralizing 17b antibody
    • Complex: HIV-1 Env BG505 SOSIP.664
      • Protein or peptide: HIV-1 Env BG505 SOSIP.664 gp41
      • Protein or peptide: HIV-1 Env BG505 SOSIP.664 gp120
    • Complex: CD4 (2-domain)
      • Protein or peptide: soluble CD4 (2-domain)
    • Complex: broadly neutralizing antibody (Fab) 17b
      • Protein or peptide: broadly neutralizing antibody (Fab) 17b light chain
      • Protein or peptide: broadly neutralizing antibody (Fab) 17b heavy chain

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Supramolecule #1: HIV-1 Env BG505 SOSIP.664 in complex with soluble CD4 (2-domain) ...

SupramoleculeName: HIV-1 Env BG505 SOSIP.664 in complex with soluble CD4 (2-domain) and Fab domain of neutralizing 17b antibody
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightTheoretical: 150 KDa

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Supramolecule #2: HIV-1 Env BG505 SOSIP.664

SupramoleculeName: HIV-1 Env BG505 SOSIP.664 / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#2
Source (natural)Organism: Human immunodeficiency virus 1
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293F

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Supramolecule #3: CD4 (2-domain)

SupramoleculeName: CD4 (2-domain) / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #3
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293F

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Supramolecule #4: broadly neutralizing antibody (Fab) 17b

SupramoleculeName: broadly neutralizing antibody (Fab) 17b / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #4-#5
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293F

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Macromolecule #1: HIV-1 Env BG505 SOSIP.664 gp41

MacromoleculeName: HIV-1 Env BG505 SOSIP.664 gp41 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
AVGIGAVFLG FLGAAGSTMG AASMTLTVQA RNLLSGIVQQ QSNLLRAPEA QQHLLKLTVW GIKQLQARVL AVERYLRDQQ LLGIWGCSGK LICCTNVPWN SSWSNRNLSE IWDNMTWLQW DKEISNYTQI IYGLLEESQN QQEKNEQDLL ALD

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Macromolecule #2: HIV-1 Env BG505 SOSIP.664 gp120

MacromoleculeName: HIV-1 Env BG505 SOSIP.664 gp120 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MDAMKRGLCC VLLLCGAVFV SPSQEIHARF RRGARAENLW VTVYYGVPVW KDAETTLFCA SDAKAYETEK HNVWATHACV PTDPNPQEIH LENVTEEFNM WKNNMVEQMH TDIISLWDQS LKPCVKLTPL CVTLQCTNVT NNITDDMRGE LKNCSFNMTT ELRDKKQKVY ...String:
MDAMKRGLCC VLLLCGAVFV SPSQEIHARF RRGARAENLW VTVYYGVPVW KDAETTLFCA SDAKAYETEK HNVWATHACV PTDPNPQEIH LENVTEEFNM WKNNMVEQMH TDIISLWDQS LKPCVKLTPL CVTLQCTNVT NNITDDMRGE LKNCSFNMTT ELRDKKQKVY SLFYRLDVVQ INENQGNRSN NSNKEYRLIN CNTSAITQAC PKVSFEPIPI HYCAPAGFAI LKCKDKKFNG TGPCPSVSTV QCTHGIKPVV STQLLLNGSL AEEEVMIRSE NITNNAKNIL VQFNTPVQIN CTRPNNNTRK SIRIGPGQAF YATGDIIGDI RQAHCNVSKA TWNETLGKVV KQLRKHFGNN TIIRFANSSG GDLEVTTHSF NCGGEFFYCN TSGLFNSTWI SNTSVQGSNS TGSNDSITLP CRIKQIINMW QRIGQAMYAP PIQGVIRCVS NITGLILTRD GGSTNSTTET FRPGGGDMRD NWRSELYKYK VVKIEPLGVA PTRCKRRVVG RRRRRR

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Macromolecule #3: soluble CD4 (2-domain)

MacromoleculeName: soluble CD4 (2-domain) / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
KKVVLGKKGD TVELTCTASQ KKSIQFHWKN SNQIKILGNQ GSFLTKGPSK LNDRADSRRS LWDQGNFPLI IKNLKIEDSD TYICEVEDQK EEVQLLVFGL TANSDTHLLQ GQSLTLTLES PPGSSPSVQC RSPRGKNIQG GKTLSVSQLE LQDSGTWTCT VLQNQKKVEF KIDIVV

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Macromolecule #4: broadly neutralizing antibody (Fab) 17b light chain

MacromoleculeName: broadly neutralizing antibody (Fab) 17b light chain / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
ELELTQSPAT LSVSPGERAT LSCRASESVS SDLAWYQQKP GQAPRLLIYG ASTRATGVPA RFSGSGSGAE FTLTISSLQS EDFAVYYCQQ YNNWPPRYTF GQGTRLEIKR

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Macromolecule #5: broadly neutralizing antibody (Fab) 17b heavy chain

MacromoleculeName: broadly neutralizing antibody (Fab) 17b heavy chain / type: protein_or_peptide / ID: 5 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
QVQLLESGAE VKKPGSSVKV SCKASGDTFI RYSFTWVRQA PGQGLEWMGR IITILDVAHY APHLQGRVTI TADKSTSTVY LELRNLRSDD TAVYFCAGVY EGEADEGEYD NNGFLKHWGQ GTLVTVT

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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

Concentration1 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
150.0 mMNaClSodium chloridesodium chloride
50.0 mMC4H11NO3Tris
1.125 mMDDM
GridModel: C-flat / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 50 % / Chamber temperature: 277 K / Instrument: HOMEMADE PLUNGER
Details: 5 microliters of the complex was incubated with 3 microliters of a fresh 1.8 mM DDM solution. A 3 microliter aliquot of the complex was applied to a C-Flat grid (CF-2/2-4C, Electron ...Details: 5 microliters of the complex was incubated with 3 microliters of a fresh 1.8 mM DDM solution. A 3 microliter aliquot of the complex was applied to a C-Flat grid (CF-2/2-4C, Electron Microscopy Sciences, Protochips, Inc.) which had been plasma cleaned for 5 seconds using a mixture of Ar/O2 (Gatan Solarus 950 Plasma system), blotted off, and then immediately plunged into liquid ethane using a manual freeze plunger..

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated defocus max: 4.0 µm / Calibrated defocus min: 1.0 µm / Calibrated magnification: 38167 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal magnification: 22500
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 90.0 K / Max: 90.0 K
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Digitization - Sampling interval: 5.0 µm / Digitization - Frames/image: 1-28 / Number grids imaged: 1 / Average exposure time: 5.6 sec. / Average electron dose: 32.0 e/Å2
Details: Individual frames were gain-corrected, aligned, and summed using MotionCor.
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: CTFFIND (ver. 3) / Details: performed internally in Relion and Frealign
Startup modelType of model: INSILICO MODEL / In silico model: common lines model using OptiMod
Details: An initial model was generated directly from the class averages using OptiMod.
Initial angle assignmentType: PROJECTION MATCHING
Projection matching processing - Angular sampling: 7.5 degrees
Software - Name: RELION (ver. 1.3) / Details: Relion 3D classification, auto mode
Final 3D classificationNumber classes: 4 / Software - Name: FREALIGN (ver. 9.11)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: FREALIGN (ver. 9.11) / Details: Frealign 3D classification and refinement
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 8.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: FREALIGN (ver. 9.11) / Details: Resolution-limited refinement used throughout / Number images used: 5716

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