9LSJ

Cryo-EM structure of the G15C-R66C and T83C-T83C diabody complex (CitS-diabody #7-TLR3)

9LSJ の概要

• エントリーDOI: 10.2210/pdb9lsj/pdb
• EMDBエントリー: 63357
• 分子名称: Citrate/sodium symporter, Toll-like receptor 3, Diabody (CitS VH-TLR3 VL), ... (9 entities in total)
• 機能のキーワード: disulfide-bridged diabody, cryo-electron microscopy (cryo-em), small protein imaging, structural marker, antibody engineering, protein nanotechnology, structural protein, structural protein-immune system complex, structural protein/immune system
• 由来する生物種: Klebsiella pneumoniae; 詳細
• タンパク質・核酸の鎖数: 5
• 化学式量合計: 257891.94

構造登録者

Kim, S.,Kim, J.W.,Park, J.G.,Lee, S.S.,Choi, S.H.,Lee, J.-O.,Jin, M.S. (登録日: 2025-02-04, 公開日: 2025-03-12, 最終更新日: 2025-06-18)

主引用文献

Kim, S.,Kim, J.W.,Park, J.G.,Lee, S.S.,Choi, S.H.,Lee, J.O.,Jin, M.S.
Disulfide-stabilized diabodies enable near-atomic cryo-EM imaging of small proteins: A case study of the bacterial Na + /citrate symporter CitS.
Structure, 33:1088-1100.e3, 2025

PubMed Abstract: Diabodies are engineered antibody fragments with two antigen-binding Fv domains. Previously, we demonstrated that they are often highly flexible but can be rigidified by introducing a disulfide bond at the Fv interface. In this study, we explored the potential of disulfide-bridged, bispecific diabodies for near-atomic cryoelectron microscopy (cryo-EM) imaging of small proteins because they can predictably link target proteins to "structural marker" proteins. As a case study, we used the bacterial citrate transporter CitS as the target protein, and the horseshoe-shaped ectodomain of human Toll-like receptor 3 (TLR3) as the marker. We show that diabodies containing one or two disulfide bonds enabled the 3D reconstruction of CitS at resolutions of 3.3 Å and 3.1 Å, respectively. This resolution surpassed previous crystallographic results and allowed us to visualize the high-resolution structural features of the transporter. Our work expands the application of diabodies in structural biology to address a key limitation in the field.

PubMed: 40169000
DOI: 10.1016/j.str.2025.03.006

実験手法

ELECTRON MICROSCOPY (3.1 Å)