Single chain variable fragment against nicastrin inhibits the γ-secretase activity

Ikuo Hayashi, Sho Takatori, Yasuomi Urano, Hiroko Iwanari, Noriko Isoo, Satoko Osawa, Maiko A. Fukuda, Tatsuhiko Kodama, Takao Hamakubo, Tong Li, Philip C. Wong, Taisuke Tomita, Takeshi Iwatsubo

Research output: Contribution to journalArticlepeer-review

Abstract

γ-Secretase is a membrane protein complex that catalyzes intramembrane proteolysis of a variety of substrates including the amyloidβ precursor protein of Alzheimer disease. Nicastrin (NCT), a single-pass membrane glycoprotein that harbors a large extracellular domain, is an essential component of the γ-secretase complex. Here we report that overexpression of a single chain variable fragment (scFv) against NCT as an intrabody suppressed the γ-secretase activity. Biochemical analyses revealed that the scFv disrupted the proper folding and the appropriate glycosyl maturation of the endogenous NCT, which are required for the stability of the γ-secretase complex and the intrinsic proteolytic activity, respectively, implicating the dual role of NCT in the γ-secretase complex. Our results also highlight the importance of the calnexin cycle in the functional maturation of the γ-secretase complex. The engineered intrabodies may serve as rationally designed, molecular targeting tools for the discovery of novel actions of the membrane proteins.

Original languageEnglish (US)
Pages (from-to)27838-27847
Number of pages10
JournalJournal of Biological Chemistry
Volume284
Issue number41
DOIs
StatePublished - Oct 9 2009

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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