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- EMDB-23480: Cryo-EM structure of llama J3 VHH antibody in complex with HIV-1 ... -

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Basic information

Entry
Database: EMDB / ID: EMD-23480
TitleCryo-EM structure of llama J3 VHH antibody in complex with HIV-1 Env BG505 DS-SOSIP.664
Map datasharpened map
Sample
  • Complex: HIV-1 Env with llama J3 VHH
    • Protein or peptide: HIV-1 Envelope Glycoprotein BG505 SOSIP.664 gp41
    • Protein or peptide: Envelope glycoprotein gp160
    • Protein or peptide: J3 VHH
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
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 / host cell endosome membrane / clathrin-dependent endocytosis of virus by host cell / viral protein processing / fusion of virus membrane with host plasma membrane / virus-mediated perturbation of host defense response / 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 / host cell endosome membrane / clathrin-dependent endocytosis of virus by host cell / viral protein processing / fusion of virus membrane with host plasma membrane / virus-mediated perturbation of host defense response / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / host cell plasma membrane / virion membrane / structural molecule activity / identical protein binding / 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
Similarity search - Component
Biological speciesHuman immunodeficiency virus 1 / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.61 Å
AuthorsGorman J / Kwong PD
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
Other privateSimons Foundation (SF349247) United States
CitationJournal: Structure / Year: 2022
Title: Structural basis for llama nanobody recognition and neutralization of HIV-1 at the CD4-binding site.
Authors: Tongqing Zhou / Lei Chen / Jason Gorman / Shuishu Wang / Young D Kwon / Bob C Lin / Mark K Louder / Reda Rawi / Erik-Stephane D Stancofski / Yongping Yang / Baoshan Zhang / Anna Forsman ...Authors: Tongqing Zhou / Lei Chen / Jason Gorman / Shuishu Wang / Young D Kwon / Bob C Lin / Mark K Louder / Reda Rawi / Erik-Stephane D Stancofski / Yongping Yang / Baoshan Zhang / Anna Forsman Quigley / Laura E McCoy / Lucy Rutten / Theo Verrips / Robin A Weiss / / Nicole A Doria-Rose / Lawrence Shapiro / Peter D Kwong /
Abstract: Nanobodies can achieve remarkable neutralization of genetically diverse pathogens, including HIV-1. To gain insight into their recognition, we determined crystal structures of four llama nanobodies ...Nanobodies can achieve remarkable neutralization of genetically diverse pathogens, including HIV-1. To gain insight into their recognition, we determined crystal structures of four llama nanobodies (J3, A12, C8, and D7), all of which targeted the CD4-binding site, in complex with the HIV-1 envelope (Env) gp120 core, and determined a cryoelectron microscopy (cryo-EM) structure of J3 with the Env trimer. Crystal and cryo-EM structures of J3 complexes revealed this nanobody to mimic binding to the prefusion-closed trimer for the primary site of CD4 recognition as well as a secondary quaternary site. In contrast, crystal structures of A12, C8, and D7 with gp120 revealed epitopes that included portions of the gp120 inner domain, inaccessible on the prefusion-closed trimer. Overall, these structures explain the broad and potent neutralization of J3 and limited neutralization of A12, C8, and D7, which utilized binding modes incompatible with the neutralization-targeted prefusion-closed conformation of Env.
History
DepositionFeb 12, 2021-
Header (metadata) releaseFeb 23, 2022-
Map releaseFeb 23, 2022-
UpdateMar 29, 2023-
Current statusMar 29, 2023Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7lpn
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23480.png

Downloads & links

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Map

FileDownload / File: emd_23480.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationsharpened map
Voxel sizeX=Y=Z: 1.0605 Å
Density
Contour LevelBy AUTHOR: 0.5 / Movie #1: 0.7
Minimum - Maximum-1.5800198 - 3.091492
Average (Standard dev.)0.015769346 (±0.106823355)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 296.94 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.06051.06051.0605
M x/y/z280280280
origin x/y/z0.0000.0000.000
length x/y/z296.940296.940296.940
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS280280280
D min/max/mean-1.5803.0910.016

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Supplemental data

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Mask #1

Fileemd_23480_msk_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Unsharpened map

Fileemd_23480_additional_1.map
AnnotationUnsharpened map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_23480_half_map_1.map
Annotationhalf map B
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Half map: Half map A

Fileemd_23480_half_map_2.map
AnnotationHalf map A
Projections & Slices
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Sample components

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Entire : HIV-1 Env with llama J3 VHH

EntireName: HIV-1 Env with llama J3 VHH
Components
  • Complex: HIV-1 Env with llama J3 VHH
    • Protein or peptide: HIV-1 Envelope Glycoprotein BG505 SOSIP.664 gp41
    • Protein or peptide: Envelope glycoprotein gp160
    • Protein or peptide: J3 VHH
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: HIV-1 Env with llama J3 VHH

SupramoleculeName: HIV-1 Env with llama J3 VHH / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Human immunodeficiency virus 1

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Macromolecule #1: HIV-1 Envelope Glycoprotein BG505 SOSIP.664 gp41

MacromoleculeName: HIV-1 Envelope Glycoprotein BG505 SOSIP.664 gp41 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightTheoretical: 17.146482 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
AVGIGAVFLG FLGAAGSTMG AASMTLTVQA RNLLSGIVQQ QSNLLRAPEA QQHLLKLTVW GIKQLQARVL AVERYLRDQQ LLGIWGCSG KLICCTNVPW NSSWSNRNLS EIWDNMTWLQ WDKEISNYTQ IIYGLLEESQ NQQEKNEQDL LALD

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Macromolecule #2: Envelope glycoprotein gp160

MacromoleculeName: Envelope glycoprotein gp160 / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightTheoretical: 53.300348 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: AENLWVTVYY GVPVWKDAET TLFCASDAKA YETEKHNVWA THACVPTDPN PQEIHLENVT EEFNMWKNNM VEQMHTDIIS LWDQSLKPC VKLTPLCVTL QCTNVTNNIT DDMRGELKNC SFNMTTELRD KKQKVYSLFY RLDVVQINEN QGNRSNNSNK E YRLINCNT ...String:
AENLWVTVYY GVPVWKDAET TLFCASDAKA YETEKHNVWA THACVPTDPN PQEIHLENVT EEFNMWKNNM VEQMHTDIIS LWDQSLKPC VKLTPLCVTL QCTNVTNNIT DDMRGELKNC SFNMTTELRD KKQKVYSLFY RLDVVQINEN QGNRSNNSNK E YRLINCNT SACTQACPKV SFEPIPIHYC APAGFAILKC KDKKFNGTGP CPSVSTVQCT HGIKPVVSTQ LLLNGSLAEE EV MIRSENI TNNAKNILVQ FNTPVQINCT RPNNNTRKSI RIGPGQAFYA TGDIIGDIRQ AHCNVSKATW NETLGKVVKQ LRK HFGNNT IIRFANSSGG DLEVTTHSFN CGGEFFYCNT SGLFNSTWIS NTSVQGSNST GSNDSITLPC RIKQIINMWQ RIGQ CMYAP PIQGVIRCVS NITGLILTRD GGSTNSTTET FRPGGGDMRD NWRSELYKYK VVKIEPLGVA PTRCKRRVVG R

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Macromolecule #3: J3 VHH

MacromoleculeName: J3 VHH / type: protein_or_peptide / ID: 3 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 14.489186 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
EVQLVESGGG LVQAGGFLRL SCELRGSIFN QYAMAWFRQA PGKEREFVAG MGAVPHYGEF VKGRFTISRD NAKSTVYLQM SSLKPEDTA IYFCARSKST YISYNSNGYD YWGRGTQVTV SSAAAHHHHH H

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Macromolecule #8: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 8 / Number of copies: 33 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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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

Concentration1.0 mg/mL
BufferpH: 7.4 / Component - Formula: PBS
GridModel: C-flat-1.2/1.3 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Average exposure time: 10.0 sec. / Average electron dose: 77.72 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 153590
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.61 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.15) / Number images used: 25685
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final 3D classificationSoftware - Name: cryoSPARC (ver. 2.12)

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Atomic model buiding 1

Initial modelPDB ID:

6nnf

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-7lpn:
Cryo-EM structure of llama J3 VHH antibody in complex with HIV-1 Env BG505 DS-SOSIP.664

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