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Yorodumi- EMDB-6099: Exploring IgG1 Antibody Conformational Flexibility in Three Dimensions -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-6099 | |||||||||
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Title | Exploring IgG1 Antibody Conformational Flexibility in Three Dimensions | |||||||||
Map data | Tomographic reconstruction of IgG1 antibody | |||||||||
Sample |
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Keywords | antibody dynamics / structural equilibrium / fluctuation / electron microscopy / individual-particle electron tomography / focused electron tomographic reconstruction | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | electron tomography / negative staining / Resolution: 20.0 Å | |||||||||
Authors | Zhang X / Zhang L / Tong H / Peng B / Rames MJ / Zhang S / Ren G | |||||||||
Citation | Journal: Sci Rep / Year: 2015 Title: 3D Structural Fluctuation of IgG1 Antibody Revealed by Individual Particle Electron Tomography. Authors: Xing Zhang / Lei Zhang / Huimin Tong / Bo Peng / Matthew J Rames / Shengli Zhang / Gang Ren / Abstract: Commonly used methods for determining protein structure, including X-ray crystallography and single-particle reconstruction, often provide a single and unique three-dimensional (3D) structure. ...Commonly used methods for determining protein structure, including X-ray crystallography and single-particle reconstruction, often provide a single and unique three-dimensional (3D) structure. However, in these methods, the protein dynamics and flexibility/fluctuation remain mostly unknown. Here, we utilized advances in electron tomography (ET) to study the antibody flexibility and fluctuation through structural determination of individual antibody particles rather than averaging multiple antibody particles together. Through individual-particle electron tomography (IPET) 3D reconstruction from negatively-stained ET images, we obtained 120 ab-initio 3D density maps at an intermediate resolution (~1-3 nm) from 120 individual IgG1 antibody particles. Using these maps as a constraint, we derived 120 conformations of the antibody via structural flexible docking of the crystal structure to these maps by targeted molecular dynamics simulations. Statistical analysis of the various conformations disclosed the antibody 3D conformational flexibility through the distribution of its domain distances and orientations. This blueprint approach, if extended to other flexible proteins, may serve as a useful methodology towards understanding protein dynamics and functions. | |||||||||
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
-Related structure data
Similar structure data |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_6099.map.gz / Format: CCP4 / Size: 39.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Tomographic reconstruction of IgG1 antibody | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.48 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
+Supplemental map: submit-120-p002.map
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-Sample components
-Entire : Human IgG1
Entire | Name: Human IgG1Immunoglobulin G |
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Components |
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-Supramolecule #1000: Human IgG1
Supramolecule | Name: Human IgG1 / type: sample / ID: 1000 / Oligomeric state: monomer / Number unique components: 1 |
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Molecular weight | Experimental: 150 KDa / Theoretical: 150 KDa |
-Macromolecule #1: immunoglobulin G subclass 1
Macromolecule | Name: immunoglobulin G subclass 1 / type: protein_or_peptide / ID: 1 / Name.synonym: IgG1 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Homo sapiens (human) / synonym: human |
Molecular weight | Experimental: 150 KDa / Theoretical: 150 KDa |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | electron tomography |
Aggregation state | particle |
-Sample preparation
Concentration | 0.01 mg/mL |
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Buffer | pH: 7.4 Details: 1X Dulbeccos phosphate-buffered saline (Invitrogen, La Jolla, CA), 2.7 mM KCl, 1.46 mM KH2PO4, 136.9 mM NaCl, 8.1 mM Na2HPO4 |
Staining | Type: NEGATIVE Details: Antibody was diluted to 0.01 mg/mL with deionized water. Aliquots of about 3uL were applied to grids. Grids were washed with deionized water three times, then washed with 1% uranyl formate ...Details: Antibody was diluted to 0.01 mg/mL with deionized water. Aliquots of about 3uL were applied to grids. Grids were washed with deionized water three times, then washed with 1% uranyl formate three times before being blotting to dryness [optimized negative-staining EM specimen preparation protocol as described in Zhang L. and Ren G., Journal of Lipid Research, (2010) 51, 1228-1236; (2011) 52, 175-84; and BBA (2013) 1830, 2150-9.] |
Grid | Details: 200 mesh glow-discharged thin carbon-coated EM grids (Cu-200CN, Pacific Grid-Tech, USA) |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
-Electron microscopy
Microscope | ZEISS LIBRA120PLUS |
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Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Calibrated magnification: 80000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 80000 |
Sample stage | Specimen holder model: OTHER / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 ° / Tilt series - Axis1 - Angle increment: 1.5 ° |
Temperature | Min: 280 K / Max: 300 K |
Date | Feb 1, 2014 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 81 / Average electron dose: 250 e/Å2 / Bits/pixel: 16 |
-Image processing
CTF correction | Details: each tilt image |
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Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 20.0 Å / Resolution method: OTHER / Software - Name: Spider, EMAN1, EMAN2 / Details: Each particle was reconstructed individually. / Number images used: 81 |
Details | Maps were reconstructed using individual-particle electron tomography (IPET) and the Focus ET Reconstruction Algorithm.{eulerAnglesDetails}: theta -60 to 60 degrees |
-Atomic model buiding 1
Initial model | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B / Chain - #2 - Chain ID: C / Chain - #3 - Chain ID: D |
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Software | Name: Chimera, NAMD |
Details | The new structure was obtained by target molecular dynamics simulation using rigid body docking results. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation |