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 language||English (US)|
|Number of pages||10|
|Journal||Journal of Biological Chemistry|
|State||Published - Oct 9 2009|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology