In vitro gamma-secretase cleavage of the Alzheimer's amyloid precursor protein correlates to a subset of presenilin complexes and is inhibited by zinc.
FEBS J
; 272(21): 5544-57, 2005 Nov.
Article
em En
| MEDLINE
| ID: mdl-16262694
The gamma-secretase complex mediates the final proteolytic event in Alzheimer's disease amyloid-beta biogenesis. This membrane complex of presenilin, anterior pharynx defective, nicastrin, and presenilin enhancer-2 cleaves the C-terminal 99-amino acid fragment of the amyloid precursor protein intramembranously at gamma-sites to form C-terminally heterogeneous amyloid-beta and cleaves at an epsilon-site to release the intracellular domain or epsilon-C-terminal fragment. In this work, two novel in vitro gamma-secretase assays are developed to further explore the biochemical characteristics of gamma-secretase activity. During development of a bacterial expression system for a substrate based on the amyloid precursor protein C-terminal 99-amino acid sequence, fragments similar to amyloid-beta and an epsilon-C-terminal fragment were observed. Upon purification this substrate was used in parallel with a transfected source of substrate to measure gamma-secretase activity from detergent extracted membranes. With these systems, it was determined that recovery of size-fractionated cellular and tissue-derived gamma-secretase activity is dependent upon detergent concentration and that activity correlates to a subset of high molecular mass presenilin complexes. We also show that by changing the solvent environment with dimethyl sulfoxide, detection of epsilon-C-terminal fragments can be elevated. Lastly, we show that zinc causes an increase in the apparent molecular mass of an amyloid precursor protein gamma-secretase substrate and inhibits its cleavage. These studies further refine our knowledge of the complexes and biochemical factors needed for gamma-secretase activity and suggest a mechanism by which zinc dysregulation may contribute to Alzheimer's disease pathogenesis.
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Base de dados:
MEDLINE
Assunto principal:
Endopeptidases
/
Zinco
/
Processamento de Proteína Pós-Traducional
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Precursor de Proteína beta-Amiloide
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Complexos Multiproteicos
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Proteínas de Membrana
Limite:
Animals
Idioma:
En
Ano de publicação:
2005
Tipo de documento:
Article