7RXC

CryoEM structure of KDELR with Legobody

Summary for 7RXC

• Entry DOI: 10.2210/pdb7rxc/pdb
• Related: 7R9D
• EMDB information: 24728
• Related PRD ID: PRD_900001
• Descriptor: Fab_8D3_2 heavy chain, Fab_8D3_2 light chain, Nb_KR, ... (7 entities in total)
• Functional Keywords: protein a, maltose-binding protein, fab, nanobody, structural protein
• Biological source: Mus musculus (mouse); More
• Total number of polymer chains: 5
• Total formula weight: 156746.14

Authors

Wu, X.D.,Rapoport, T.A. (deposition date: 2021-08-22, release date: 2021-10-06, Last modification date: 2025-06-04)

Primary citation

Wu, X.,Rapoport, T.A.
Cryo-EM structure determination of small proteins by nanobody-binding scaffolds (Legobodies).
Proc.Natl.Acad.Sci.USA, 118:-, 2021

PubMed 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 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.

PubMed: 34620716
DOI: 10.1073/pnas.2115001118

Experimental method

ELECTRON MICROSCOPY (3.2 Å)