+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-6296 | |||||||||
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Title | Bg505 gp140 NFL2P liganded to PGV04 | |||||||||
Map data | Bg505 gp140 NFL2 liganded to PGV04 | |||||||||
Sample |
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Biological species | Human immunodeficiency virus 1 / Homo sapiens (human) | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 17.0 Å | |||||||||
Authors | de Val N / Sharma S / Bale S | |||||||||
Citation | Journal: Cell Rep / Year: 2015 Title: Cleavage-independent HIV-1 Env trimers engineered as soluble native spike mimetics for vaccine design. Authors: Shailendra Kumar Sharma / Natalia de Val / Shridhar Bale / Javier Guenaga / Karen Tran / Yu Feng / Viktoriya Dubrovskaya / Andrew B Ward / Richard T Wyatt / Abstract: Viral glycoproteins mediate entry by pH-activated or receptor-engaged activation and exist in metastable pre-fusogenic states that may be stabilized by directed rational design. As recently reported, ...Viral glycoproteins mediate entry by pH-activated or receptor-engaged activation and exist in metastable pre-fusogenic states that may be stabilized by directed rational design. As recently reported, the conformationally fixed HIV-1 envelope glycoprotein (Env) trimers in the pre-fusion state (SOSIP) display molecular homogeneity and structural integrity at relatively high levels of resolution. However, the SOSIPs necessitate full Env precursor cleavage, which requires endogenous furin overexpression. Here, we developed an alternative strategy using flexible peptide covalent linkage of Env subdomains to produce soluble, homogeneous, and cleavage-independent Env mimics, called native flexibly linked (NFL) trimers, as vaccine candidates. This simplified design avoids the need for furin co-expression and, in one case, antibody affinity purification to accelerate trimer scale-up for preclinical and clinical applications. We have successfully translated the NFL design to multiple HIV-1 subtypes, establishing the potential to become a general method of producing native-like, well-ordered Env trimers for HIV-1 or other viruses. | |||||||||
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_6296.map.gz | 14.8 MB | EMDB map data format | |
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Header (meta data) | emd-6296-v30.xml emd-6296.xml | 10 KB 10 KB | Display Display | EMDB header |
Images | 400_6296.gif 80_6296.gif | 13 KB 1.8 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-6296 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-6296 | HTTPS FTP |
-Validation report
Summary document | emd_6296_validation.pdf.gz | 77.6 KB | Display | EMDB validaton report |
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Full document | emd_6296_full_validation.pdf.gz | 76.7 KB | Display | |
Data in XML | emd_6296_validation.xml.gz | 494 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6296 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6296 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_6296.map.gz / Format: CCP4 / Size: 15.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Bg505 gp140 NFL2 liganded to PGV04 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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 : Bg505 gp140 NFL liganded to PGV04
Entire | Name: Bg505 gp140 NFL liganded to PGV04 |
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Components |
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-Supramolecule #1000: Bg505 gp140 NFL liganded to PGV04
Supramolecule | Name: Bg505 gp140 NFL liganded to PGV04 / type: sample / ID: 1000 / Oligomeric state: trimer / Number unique components: 2 |
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Molecular weight | Experimental: 570 KDa / Theoretical: 570 KDa / Method: Size exclusion chromatography (SEC) |
-Macromolecule #1: Bg505 gp140 NFL2P
Macromolecule | Name: Bg505 gp140 NFL2P / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Oligomeric state: trimer / Recombinant expression: Yes |
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Source (natural) | Organism: Human immunodeficiency virus 1 / synonym: HIV-1 |
Molecular weight | Experimental: 570 KDa / Theoretical: 570 KDa |
Recombinant expression | Organism: Homo sapiens (human) / Recombinant cell: HEK 293T |
-Macromolecule #2: PGV04 Fab
Macromolecule | Name: PGV04 Fab / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Homo sapiens (human) / synonym: human |
-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 / Details: 50 mM Tris-HCl, 150 mM NaCl |
Staining | Type: NEGATIVE / Details: 2% uranyl formate for 30 seconds |
Grid | Details: 400 mesh Cu grids, negatively glow discharged for 30 seconds |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
-Electron microscopy
Microscope | FEI TECNAI SPIRIT |
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Date | May 16, 2014 |
Image recording | Category: CCD / Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Number real images: 131 / Average electron dose: 40.30 e/Å2 |
Tilt angle min | 0 |
Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Calibrated magnification: 52000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 46000 |
Sample stage | Specimen holder model: HOME BUILD / Tilt angle max: 50 |
Experimental equipment | Model: Tecnai Spirit / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 17.0 Å / Resolution method: OTHER / Software - Name: EMAN, sparx / Number images used: 28143 |
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-Atomic model buiding 1
Initial model | PDB ID: |
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Refinement | Space: REAL / Protocol: RIGID BODY FIT |