Presenilins are polytopic, integral proteins that control intramembranous proteolysis at the "γ-" and "ε-" cleavage sites of the Alzheimer amyloid-β precursor protein (APP) to yield amyloid-β peptide (Aβ) and the APP intracellular domain (AICD). We have overexpressed a constitutively active, pathogenic form of PS1 (known as PS1 Δ exon 9) together with its substrate, APP-C99, in Spodoperta frugiperda (Sf9) cells. Sf9 cells have been reported to lack endogenous γ-secretase, an unexpected finding since there exists an insect homologue of PS1. In our hands, neither intact insect cells coexpressing PS1 Δ exon 9/APP-C99 nor the aqueous homogenates of these cells displayed obvious products of the γ- or ε-secretase reactions, as reported. Surprisingly, when APP-C99-expressing cells were homogenized in 3[(3-cholamidopropyl) dimethylammonio]-2- hydroxypropanesulfonic acid (CHAPSO), a detergent known to support γ-secretase activity, subsequent incubation led to the accumulation of an AICD-like peptide (AICD-L). Aspartyl proteinase inhibitors were effective in preventing the appearance of AICD-L, but inhibitors of other classes of proteinases were ineffective. Immunoprecipitation-mass spectrometry of AICD-L revealed its identity as the minor of the two known AICDs.
|Original language||English (US)|
|Number of pages||3|
|Journal||Alzheimer disease and associated disorders|
|State||Published - Oct 2004|