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Yorodumi- EMDB-22999: Negative stain reconstruction of murine antibody M4H2K1 Fab in co... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-22999 | |||||||||
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Title | Negative stain reconstruction of murine antibody M4H2K1 Fab in complex with HIV-1 Env BG505 UFO.664 trimer | |||||||||
Map data | full map | |||||||||
Sample |
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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 / apoptotic process / host cell plasma membrane / virion membrane / structural molecule activity / plasma membrane Similarity search - Function | |||||||||
Biological species | Mus musculus (house mouse) | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 25.0 Å | |||||||||
Authors | Copps J / Ozorowski G / Ward AB | |||||||||
Funding support | United States, 1 items
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Citation | Journal: mBio / Year: 2021 Title: Neutralizing Antibodies Induced by First-Generation gp41-Stabilized HIV-1 Envelope Trimers and Nanoparticles. Authors: Sonu Kumar / Xiaohe Lin / Timothy Ngo / Benjamin Shapero / Cindy Sou / Joel D Allen / Jeffrey Copps / Lei Zhang / Gabriel Ozorowski / Linling He / Max Crispin / Andrew B Ward / Ian A Wilson / Jiang Zhu / Abstract: The immunogenicity of gp41-stabilized HIV-1 BG505 envelope (Env) trimers and nanoparticles (NPs) was recently assessed in mice and rabbits. Here, we combined Env-specific B-cell sorting and ...The immunogenicity of gp41-stabilized HIV-1 BG505 envelope (Env) trimers and nanoparticles (NPs) was recently assessed in mice and rabbits. Here, we combined Env-specific B-cell sorting and repertoire sequencing to identify neutralizing antibodies (NAbs) from immunized animals. A panel of mouse NAbs was isolated from mice immunized with a 60-meric I3-01 NP presenting 20 stabilized trimers. Three mouse NAbs potently neutralized BG505.T332N by recognizing a glycan epitope centered in the C3/V4 region on BG505 Env, as revealed by electron microscopy (EM), X-ray crystallography, and epitope mapping. A set of rabbit NAbs was isolated from rabbits immunized with a soluble trimer and a 24-meric ferritin NP presenting 8 trimers. Neutralization assays against BG505.T332N variants confirmed that potent rabbit NAbs targeted previously described glycan holes on BG505 Env and accounted for a significant portion of the autologous NAb response in both the trimer and ferritin NP groups. Last, we examined NAb responses that were induced by non-BG505 Env immunogens. We determined a 3.4-Å-resolution crystal structure for the clade C transmitted/founder (T/F) Du172.17 Env with a redesigned heptad repeat 1 (HR1) bend in gp41. This clade C Env, in a soluble trimer form and in a multivalent form with 8 trimers attached to ferritin NP, and the gp41-stabilized clade A Q482-d12 Env trimer elicited distinct NAb responses in rabbits, with notable differences in neutralization breadth. Although eliciting a broad NAb response remains a major challenge, our study provides valuable information on an HIV-1 vaccine design strategy that combines gp41 stabilization and NP display. Self-assembling protein nanoparticles (NPs) presenting BG505 envelope (Env) trimers can elicit tier 2 HIV-1-neutralizing antibody (NAb) responses more effectively than soluble trimers. In the present study, monoclonal NAbs were isolated from previously immunized mice and rabbits for structural and functional analyses, which revealed that potent mouse NAbs recognize the C3/V4 region and small NP-elicited rabbit NAbs primarily target known glycan holes on BG505 Env. This study validates the gp41 stabilization strategy for HIV-1 Env vaccine design and highlights the challenge in eliciting a broad NAb response. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_22999.map.gz | 7.6 MB | EMDB map data format | |
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Header (meta data) | emd-22999-v30.xml emd-22999.xml | 10.1 KB 10.1 KB | Display Display | EMDB header |
Images | emd_22999.png | 58 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-22999 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-22999 | HTTPS FTP |
-Validation report
Summary document | emd_22999_validation.pdf.gz | 288.5 KB | Display | EMDB validaton report |
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Full document | emd_22999_full_validation.pdf.gz | 288.1 KB | Display | |
Data in XML | emd_22999_validation.xml.gz | 5.7 KB | Display | |
Data in CIF | emd_22999_validation.cif.gz | 6.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-22999 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-22999 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_22999.map.gz / Format: CCP4 / Size: 15.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | full map | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.05 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : 3 copies of M4H2K1 Fab in complex with trimeric HIV-1 Env BG505 U...
Entire | Name: 3 copies of M4H2K1 Fab in complex with trimeric HIV-1 Env BG505 UFO.664 |
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Components |
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-Supramolecule #1: 3 copies of M4H2K1 Fab in complex with trimeric HIV-1 Env BG505 U...
Supramolecule | Name: 3 copies of M4H2K1 Fab in complex with trimeric HIV-1 Env BG505 UFO.664 type: complex / ID: 1 / Parent: 0 |
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Source (natural) | Organism: Mus musculus (house mouse) |
Recombinant expression | Organism: Homo sapiens (human) / Recombinant strain: HEK293F |
Molecular weight | Theoretical: 570 KDa |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.01 mg/mL | ||||||
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Buffer | pH: 7.4 Component:
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Staining | Type: NEGATIVE / Material: uranyl formate / Details: 2% (w/v) UF applied for 60 s | ||||||
Grid | Model: Homemade / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: PLASMA CLEANING |
-Electron microscopy
Microscope | FEI TECNAI SPIRIT |
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Image recording | Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Average electron dose: 25.0 e/Å2 |
Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal magnification: 52000 |
Sample stage | Specimen holder model: SIDE ENTRY, EUCENTRIC |
Experimental equipment | Model: Tecnai Spirit / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: PDB ENTRY PDB model - PDB ID: Details: Map created from PDB coordinates and low pass filtered |
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Final reconstruction | Applied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 25.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: cryoSPARC (ver. v2) / Number images used: 14027 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |