8VGO

CryoEM structure of CD20 in complex with engineered conformationally rigid Rituximab.4DS Fab

8VGO の概要

• エントリーDOI: 10.2210/pdb8vgo/pdb
• 関連するPDBエントリー: 8VEG 8VGE 8VGF 8VGG 8VGH 8VGI 8VGJ 8VGK 8VGL 8VGM 8VGN
• EMDBエントリー: 43200 43201 43202 43203 43204 43205 43206 43207 43208 43209 43210 43211 43212 43213 43214 43215 43216
• 分子名称: Rituximab.4DS Fab heavy chain, Rituximab.4DS Fab light chain, B-lymphocyte antigen CD20 (3 entities in total)
• 機能のキーワード: antibody fragment, fab, protein engineering, membrane protein, membrane protein-immune system complex, membrane protein/immune system
• 由来する生物種: Homo sapiens; 詳細
• タンパク質・核酸の鎖数: 6
• 化学式量合計: 159545.91

構造登録者

Kung, J.E.,Jao, C.C.,Arthur, C.P.,Sudhamsu, J. (登録日: 2023-12-27, 公開日: 2024-11-06, 最終更新日: 2024-11-27)

主引用文献

Kung, J.E.,Johnson, M.C.,Jao, C.C.,Arthur, C.P.,Tegunov, D.,Rohou, A.,Sudhamsu, J.
Disulfide constrained Fabs overcome target size limitation for high-resolution single-particle cryo-EM.
Biorxiv, 2024

PubMed Abstract: High-resolution structures of proteins are critical to understanding molecular mechanisms of biological processes and in the discovery of therapeutic molecules. Cryo-EM has revolutionized structure determination of large proteins and their complexes, but a vast majority of proteins that underlie human diseases are small (< 50 kDa) and usually beyond its reach due to low signal-to-noise images and difficulties in particle alignment. Current strategies to overcome this problem increase the overall size of small protein targets using scaffold proteins that bind to the target, but are limited by inherent flexibility and not being bound to their targets in a rigid manner, resulting in the target being poorly resolved compared to the scaffolds. Here we present an iteratively engineered molecular design for transforming Fabs (antibody fragments), into conformationally rigid scaffolds (Rigid-Fabs) that, when bound to small proteins (~20 kDa), can enable high-resolution structure determination using cryo-EM. This design introduces multiple disulfide bonds at strategic locations, generates a well-folded Fab constrained into a rigid conformation and can be applied to Fabs from various species, isotypes and chimeric Fabs. We present examples of the Rigid Fab design enabling high-resolution (2.3-2.5 Å) structures of small proteins, Ang2 (26 kDa) and KRAS (21 kDa) by cryo-EM. The strategies for designing disulfide constrained Rigid Fabs in our work thus establish a general approach to overcome the target size limitation of single particle cryo-EM.

PubMed: 38798381
DOI: 10.1101/2024.05.10.593593

実験手法

ELECTRON MICROSCOPY (2.6 Å)