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Alzheimer amyloid precursor aspartyl proteinase activity in CHAPSO homogenates of Spodoptera frugiperda cells


Presenilins are polytopic, integral proteins that control intramembranous proteolysis at the ""gamma-"" and ""epsilon-"" cleavage sites of the Alzheimer amyloid-beta precursor protein (APP) to yield amyloid-beta peptide (Abeta) and the APP intracellular domain (AICD). We have overexpressed a constitutively active, pathogenic form of PS1 (known as PS1 Delta exon 9) together with its substrate, APP-C99, in Spodoptera frugiperda (Sf9) cells. Sf9 cells have been reported to lack endogenous gamma-secretase, an unexpected finding since there exists an insect homologue of PS1. In our hands, neither intact insect cells coexpressing PS1 Delta exon 9/APP-C99 nor the aqueous homogenates of these cells displayed obvious products of the gamma- or epsilon-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 gamma-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.

Type Journal
ISBN 0893-0341 (Print)
Authors Carter, T. L.;Verdile, G.;Groth, D.;Bogush, A.;Thomas, S.;Shen, P.;Fraser, P. E.;Mathews, P.;Nixon, R. A.;Ehrlich, M. E.;Kwok, J. B.;St George-Hyslop, P.;Schofield, P.;Li, Y.;Yang, A.;Martins, R. N.;Gandy, S. :
Published Date 2004-01-01
Published Volume 18
Published Issue 4
Published Pages 261-3
Status Published in-print
URL link to publisher's version