Assays for amyloid precursor protein α -secretase activity

William A. Campbell, Michael S. Wolfe, Weiming Xia

Research output: Chapter in Book/Report/Conference proceedingChapter

1 Scopus citations

Abstract

α -Secretase cleavage, mediated by a complex of presenilin (PS), nicastrin, PEN-2, and APH-1, is the final proteolytic step in generating amyloid beta (A β) protein and the Notch intracellular domain. A β and Notch are critical in the pathogenesis of Alzheimer’s disease (AD) and in development, respectively. In addition to cleaving amyloid precursor protein (APP) and Notch, α -secretase also cleaves over a dozen additional type I transmembrane domain proteins. α -Secretase activity can be measured in vivo by collecting conditioned media from tissue cultured cells and in vitroby incubating endogenous substrate with total microsomes or with Golgi/trans-Golgi network (TGN)-enriched microsomal vesicles or incubating recombinant substrate with solubilized membranes. For APP, Western blotting or enzyme-linked immunosorbent assay (ELISA) has been used to quantify the generation of A β 40, A β 42, and amyloid intracellular domain (AICD). These methods can be applied to study the α -secretase cleavage of any α -secretase substrate in a variety of experimental conditions.

Original languageEnglish (US)
Title of host publicationAmyloid Precursor Protein
Subtitle of host publicationA Practical Approach
PublisherCRC Press
Pages51-68
Number of pages18
ISBN (Electronic)9780203492185
ISBN (Print)9780849322457
StatePublished - Jan 1 2004

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Neuroscience(all)

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