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Open data
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Basic information
Entry | Database: PDB / ID: 7rxc | ||||||
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Title | CryoEM structure of KDELR with Legobody | ||||||
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![]() | STRUCTURAL PROTEIN / protein A / maltose-binding protein / Fab / nanobody | ||||||
Function / homology | ![]() KDEL sequence binding / COPI-dependent Golgi-to-ER retrograde traffic / COPI-coated vesicle membrane / ER retention sequence binding / protein retention in ER lumen / COPI-mediated anterograde transport / maintenance of protein localization in endoplasmic reticulum / cis-Golgi network / retrograde vesicle-mediated transport, Golgi to endoplasmic reticulum / IgG binding ...KDEL sequence binding / COPI-dependent Golgi-to-ER retrograde traffic / COPI-coated vesicle membrane / ER retention sequence binding / protein retention in ER lumen / COPI-mediated anterograde transport / maintenance of protein localization in endoplasmic reticulum / cis-Golgi network / retrograde vesicle-mediated transport, Golgi to endoplasmic reticulum / IgG binding / carbohydrate transmembrane transporter activity / endoplasmic reticulum to Golgi vesicle-mediated transport / protein transport / periplasmic space / Golgi membrane / endoplasmic reticulum membrane / endoplasmic reticulum / extracellular region / membrane Similarity search - Function | ||||||
Biological species | ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||
![]() | Wu, X.D. / Rapoport, T.A. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Cryo-EM structure determination of small proteins by nanobody-binding scaffolds (Legobodies). Authors: Xudong Wu / Tom A Rapoport / ![]() Abstract: We describe a general method that allows structure determination of small proteins by single-particle cryo-electron microscopy (cryo-EM). The method is based on the availability of a target-binding ...We describe a general method that allows structure determination of small proteins by single-particle cryo-electron microscopy (cryo-EM). The method is based on the availability of a target-binding nanobody, which is then rigidly attached to two scaffolds: 1) a Fab fragment of an antibody directed against the nanobody and 2) a nanobody-binding protein A fragment fused to maltose binding protein and Fab-binding domains. The overall ensemble of ∼120 kDa, called Legobody, does not perturb the nanobody-target interaction, is easily recognizable in EM images due to its unique shape, and facilitates particle alignment in cryo-EM image processing. The utility of the method is demonstrated for the KDEL receptor, a 23-kDa membrane protein, resulting in a map at 3.2-Å overall resolution with density sufficient for de novo model building, and for the 22-kDa receptor-binding domain (RBD) of SARS-CoV-2 spike protein, resulting in a map at 3.6-Å resolution that allows analysis of the binding interface to the nanobody. The Legobody approach thus overcomes the current size limitations of cryo-EM analysis. | ||||||
History |
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Structure visualization
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 219.6 KB | Display | ![]() |
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PDB format | ![]() | 176.8 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 24728MC ![]() 7r9dC ![]() 7rxdC C: citing same article ( M: map data used to model this data |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
-Antibody , 4 types, 4 molecules HLNB
#1: Antibody | Mass: 25252.217 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
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#2: Antibody | Mass: 24095.852 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
#3: Antibody | Mass: 14698.260 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
#4: Antibody | Mass: 59233.246 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() ![]() Gene: DAH37_23060, spa, spg / Production host: ![]() ![]() References: UniProt: A0A4Z0THX4, UniProt: P99134, UniProt: P06654 |
-Protein / Sugars / Non-polymers , 3 types, 6 molecules K![](data/chem/img/POV.gif)
![](data/chem/img/POV.gif)
#5: Protein | Mass: 30083.959 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
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#6: Polysaccharide | alpha-D-glucopyranose-(1-4)-alpha-D-glucopyranose |
#7: Chemical | ChemComp-POV / ( |
-Details
Has ligand of interest | N |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: The complex of KDELR with Legobody / Type: COMPLEX / Entity ID: #1-#5 / Source: MULTIPLE SOURCES |
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Molecular weight | Value: 0.15 MDa / Experimental value: NO |
Buffer solution | pH: 7.4 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 51.44 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
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Processing
Software | Name: PHENIX / Version: 1.19.1_4122: / Classification: refinement | ||||||||||||||||||||||||
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 246878 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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