5I76

Crystal structure of FM318, a recombinant Fab adopted from cetuximab

5I76 の概要

• エントリーDOI: 10.2210/pdb5i76/pdb
• 分子名称: FM318_light_chain, FM318_heavy_cahin (3 entities in total)
• 機能のキーワード: antibody fragment, recombinant fab, cetuximab, egfr, immune system
• 由来する生物種: Homo sapiens/Mus musculus xenograft; 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 95185.76

構造登録者

Sim, D.W.,Kim, J.H.,Seok, S.H.,Seo, M.D.,Kim, Y.P.,Won, H.S. (登録日: 2016-02-16, 公開日: 2016-12-07, 最終更新日: 2024-10-30)

主引用文献

Kim, J.H.,Sim, D.W.,Park, D.,Jung, T.G.,Lee, S.,Oh, T.,Ha, J.R.,Seok, S.H.,Seo, M.D.,Kang, H.C.,Kim, Y.P.,Won, H.S.
Bacterial production and structure-functional validation of a recombinant antigen-binding fragment (Fab) of an anti-cancer therapeutic antibody targeting epidermal growth factor receptor.
Appl.Microbiol.Biotechnol., 100:10521-10529, 2016

PubMed Abstract: Fragment engineering of monoclonal antibodies (mAbs) has emerged as an excellent paradigm to develop highly efficient therapeutic and/or diagnostic agents. Engineered mAb fragments can be economically produced in bacterial systems using recombinant DNA technologies. In this work, we established recombinant production in Escherichia coli for monovalent antigen-binding fragment (Fab) adopted from a clinically used anticancer mAB drug cetuximab targeting epidermal growth factor receptor (EGFR). Recombinant DNA constructs were designed to express both polypeptide chains comprising Fab in a single vector and to secrete them to bacterial periplasmic space for efficient folding. Particularly, a C-terminal engineering to confer an interchain disulfide bond appeared to be able to enhance its heterodimeric integrity and EGFR-binding activity. Conformational relevance of the purified final product was validated by mass spectrometry and crystal structure at 1.9 Å resolution. Finally, our recombinant cetuximab-Fab was found to have strong binding affinity to EGFR overexpressed in human squamous carcinoma model (A431) cells. Its binding ability was comparable to that of cetuximab. Its EGFR-binding affinity was estimated at approximately 0.7 nM of Kd in vitro, which was quite stronger than the binding affinity of natural ligand EGF. Hence, the results validate that our construction could serve as an efficient platform to produce a recombinant cetuximab-Fab with a retained antigen-binding functionality.

PubMed: 27470143
DOI: 10.1007/s00253-016-7717-z

実験手法

X-RAY DIFFRACTION (1.922 Å)