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- EMDB-3120: cryoEM reconstruction of bnAb PGT128 in complex with BG505 SOSIP.... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-3120 | |||||||||
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Title | cryoEM reconstruction of bnAb PGT128 in complex with BG505 SOSIP.664 Env trimer | |||||||||
![]() | Reconstruction of PGT128 Fab in complex with BG505 SOSIP.664 Env trimer | |||||||||
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![]() | HIV-1 / Env / bnAb / antibody / PGT128 | |||||||||
Function / homology | ![]() 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 / structural molecule activity / virion membrane / identical protein binding / plasma membrane Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.47 Å | |||||||||
![]() | Lee JH / de Val N / Lyumkis D / Ward AB | |||||||||
![]() | ![]() Title: Model Building and Refinement of a Natively Glycosylated HIV-1 Env Protein by High-Resolution Cryoelectron Microscopy. Authors: Jeong Hyun Lee / Natalia de Val / Dmitry Lyumkis / Andrew B Ward / ![]() Abstract: Secretory and membrane proteins from mammalian cells undergo post-translational modifications, including N-linked glycosylation, which can result in a large number of possible glycoforms. This sample ...Secretory and membrane proteins from mammalian cells undergo post-translational modifications, including N-linked glycosylation, which can result in a large number of possible glycoforms. This sample heterogeneity can be problematic for structural studies, particularly X-ray crystallography. Thus, crystal structures of heavily glycosylated proteins such as the HIV-1 Env viral spike protein have been determined by removing the majority of glycans. This step is most frequently carried out using Endoglycosidase H (EndoH) and requires that all expressed glycans be in the high-mannose form, which is often not the native glycoform. With significantly improved technologies in single-particle cryoelectron microscopy, we demonstrate that it is now possible to refine and build natively glycosylated HIV-1 Env structures in solution to 4.36 Å resolution. At this resolution we can now analyze the complete epitope of a broadly neutralizing antibody (bnAb), PGT128, in the context of the trimer expressed with native glycans. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 59.1 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 12.3 KB 12.3 KB | Display Display | ![]() |
Images | ![]() | 117.6 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Reconstruction of PGT128 Fab in complex with BG505 SOSIP.664 Env trimer | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.31 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : PGT128 Fab bound to BG505 SOSIP.664 HIV-1 Env trimer
Entire | Name: PGT128 Fab bound to BG505 SOSIP.664 HIV-1 Env trimer |
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Components |
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-Supramolecule #1000: PGT128 Fab bound to BG505 SOSIP.664 HIV-1 Env trimer
Supramolecule | Name: PGT128 Fab bound to BG505 SOSIP.664 HIV-1 Env trimer / type: sample / ID: 1000 Oligomeric state: Three monomers of PGT128 Fab bind one Env trimer Number unique components: 2 |
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Molecular weight | Theoretical: 570 KDa |
-Macromolecule #1: HIV-1 Envelope glycoprotein
Macromolecule | Name: HIV-1 Envelope glycoprotein / type: protein_or_peptide / ID: 1 / Name.synonym: Env Details: The Env sequence is from the clade A virus BG505, truncated at residue 664 of gp41, and contains stabilizing SOSIP mutations. Number of copies: 3 / Oligomeric state: Trimer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 420 KDa |
Recombinant expression | Organism: ![]() |
-Macromolecule #2: Immunoglobulin G PGT128
Macromolecule | Name: Immunoglobulin G PGT128 / type: protein_or_peptide / ID: 2 / Name.synonym: IgG PGT128 Details: The fragment antigen binding (Fab) of PGT128 was used to form the complex. Number of copies: 3 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 500 KDa |
Recombinant expression | Organism: ![]() |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 2.5 mg/mL |
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Buffer | pH: 7.4 / Details: 50 mM Tris, 150 mM NaCl, 0.675 mM DDM |
Grid | Details: 400 mesh C-Flat CF-2/2-4C, plasma treated for 5 seconds |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 160 K / Instrument: HOMEMADE PLUNGER Method: Grids were frozen using a manual plunger at 4 degrees. |
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Electron microscopy #1
Microscopy ID | 1 |
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Microscope | FEI TITAN KRIOS |
Alignment procedure | Legacy - Astigmatism: Objective astigmatism was corrected at 22,500x magnification |
Date | Oct 7, 2014 |
Image recording | Category: CCD / Film or detector model: GATAN K2 (4k x 4k) / Digitization - Sampling interval: 5 µm / Number real images: 2111 / Average electron dose: 33 e/Å2 Details: Each full dose image is an aligned stack of frames recorded each using a dose of ~10 e-/Angstrom^2/sec. |
Tilt angle min | 0 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 22500 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 22500 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Electron microscopy #2
Microscopy ID | 2 |
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Microscope | FEI TITAN KRIOS |
Alignment procedure | Legacy - Astigmatism: Objective astigmatism was corrected at 22,500x magnification |
Date | Nov 7, 2014 |
Image recording | Category: CCD / Film or detector model: GATAN K2 (4k x 4k) / Digitization - Sampling interval: 5 µm / Number real images: 2111 / Average electron dose: 35 e/Å2 Details: Each full dose image is an aligned stack of frames recorded each using a dose of ~10 e-/Angstrom^2/sec. |
Tilt angle min | 0 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 22500 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 22500 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
CTF correction | Details: Each micrograph |
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Final reconstruction | Applied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 4.47 Å / Resolution method: OTHER / Software - Name: Imagic, Relion / Number images used: 92095 |