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Solubilization of full‐length amyloid precursor proteins from PC12 cell membranes
Author(s) -
Ripellino J. A.,
Vassilacopoulou D.,
Robakis N. K.
Publication year - 1994
Publication title -
journal of neuroscience research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.72
H-Index - 160
eISSN - 1097-4547
pISSN - 0360-4012
DOI - 10.1002/jnr.490390211
Subject(s) - leupeptin , transmembrane protein , proteases , biochemistry , chemistry , protease , amyloid precursor protein , alpha secretase , serine protease , membrane protein , membrane , enzyme , alzheimer's disease , medicine , receptor , disease , pathology
The amyloid β protein (Aβ) of Alzheimer disease (AD) is derived from the proteolytic processing of the amyloid precursor proteins (APP), which are considered type I transmembrane proteins. Production of Aβ from a tranmembrane precursor predicts a proteolytic cleavage within the lipid bilayer, a site relatively inaccessible to proteases. Here we show that incubation of a membrane fraction of PC12 cells at 37°C results in the solubilization of an APP species which migrates on SDS‐PAGE as full‐length APP. The release of this full‐length APP was pH‐dependent with a peak of activity of pH 9.0. At this pH about 19% of the membrane APP was released from the active subcellular fraction. Under the same conditions other transmembrane proteins remained insoluble. Very little APP was solubilized at 4°C. APP solubilization was specifically inhibited by the serine protease inhibitors aprotinin and pefabloc. Other protease inhibitors, including leupeptin and α1‐antitrypsin, had no effect. Several metal cations, including Ca + + and Zn + + , also inhibited release of soluble full‐length APP. Low levels of full‐length APP were also detected in both the soluble fraction of PC12 cell extracts and in the media of PC12 cell cultures. These data suggest the involvement of serine protease in the solubilization of membrane, full‐length APP. The release of this APP could provide a soluble substrate for the proteolytic enzymes involved in the production of Aβ.Copyright © 1994 Wiley‐Liss, Inc.