5XHF

Crystal structure of Trastuzumab Fab fragment bearing p-azido-L-phenylalanine

5XHF の概要

• エントリーDOI: 10.2210/pdb5xhf/pdb
• 関連するPDBエントリー: 5XHG
• 分子名称: polypeptide (L chain), polypeptide (H chain) (2 entities in total)
• 機能のキーワード: fab, immune system
• 由来する生物種: MUS MUSCULUS (MOUSE); 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 93904.55

構造登録者

Kuratani, M.,Yanagisawa, T.,Sakamoto, K.,Yokoyama, S. (登録日: 2017-04-20, 公開日: 2017-12-20, 最終更新日: 2023-11-22)

主引用文献

Kato, A.,Kuratani, M.,Yanagisawa, T.,Ohtake, K.,Hayashi, A.,Amano, Y.,Kimura, K.,Yokoyama, S.,Sakamoto, K.,Shiraishi, Y.
Extensive Survey of Antibody Invariant Positions for Efficient Chemical Conjugation Using Expanded Genetic Codes.
Bioconjug. Chem., 28:2099-2108, 2017

PubMed Abstract: The site-specific chemical conjugation of proteins, following synthesis with an expanded genetic code, promises to advance antibody-based technologies, including antibody drug conjugation and the creation of bispecific Fab dimers. The incorporation of non-natural amino acids into antibodies not only guarantees site specificity but also allows the use of bio-orthogonal chemistry. However, the efficiency of amino acid incorporation fluctuates significantly among different sites, thereby hampering the identification of useful conjugation sites. In this study, we applied the codon reassignment technology to achieve the robust and efficient synthesis of chemically functionalized antibodies containing N-(o-azidobenzyloxycarbonyl)-l-lysine (o-Az-Z-Lys) at defined positions. This lysine derivative has a bio-orthogonally reactive group at the end of a long side chain, enabling identification of multiple new positions in Fab-constant domains, allowing chemical conjugation with high efficiency. An X-ray crystallographic study of a Fab variant with o-Az-Z-Lys revealed high-level exposure of the azido group to solvent, with six of the identified positions subsequently used to engineer "Variabodies", a novel antibody format allowing various connections between two Fab molecules. Our findings indicated that some of the created Variabodies exhibited agonistic activity in cultured cells as opposed to the antagonistic nature of antibodies. These results showed that our approach greatly enhanced the availability of antibodies for chemical conjugation and might aid in the development of new therapeutic antibodies.

PubMed: 28727448
DOI: 10.1021/acs.bioconjchem.7b00265

実験手法

X-RAY DIFFRACTION (3.205 Å)