Epitope Mapping for Monoclonal Antibody Reveals the Activation Mechanism for αVβ3 Integrin

Tetsuji Kamata, Makoto Handa, Sonomi Takakuwa, Yukiko Sato, Yohko Kawai, Yasuo Ikeda, Sadakazu Aiso

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Epitopes for a panel of anti-αVβ3 monoclonal antibodies (mAbs) were investigated to explore the activation mechanism of αVβ3 integrin. Experiments utilizing αV/αIIb domain-swapping chimeras revealed that among the nine mAbs tested, five recognized the ligand-binding β-propeller domain and four recognized the thigh domain, which is the upper leg of the αV chain. Interestingly, the four mAbs included function-blocking as well as non-functional mAbs, although they bound at a distance from the ligand-binding site. The epitopes for these four mAbs were further determined using human-to-mouse αV chimeras. Among the four, P3G8 recognized an amino acid residue, Ser-528, located on the side of the thigh domain, while AMF-7, M9, and P2W7 all recognized a common epitope, Ser-462, that was located close to the α-genu, where integrin makes a sharp bend in the crystal structure. Fibrinogen binding studies for cells expressing wild-type αVβ3 confirmed that AMF-7, M9, and P2W7 were inhibitory, while P3G8 was non-functional. However, these mAbs were all unable to block binding when αVβ3 was constrained in its extended conformation. These results suggest that AMF-7, M9, and P2W7 block ligand binding allosterically by stabilizing the angle of the bend in the bent conformation. Thus, a switchblade-like movement of the integrin leg is indispensable for the affinity regulation of αVβ3 integrin.

Original languageEnglish
Article numbere66096
JournalPloS one
Volume8
Issue number6
DOIs
Publication statusPublished - 2013 Jun 20

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

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