The amyloid beta protein (A beta) 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 A beta from a transmembrane 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 degrees 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 degrees C. APP solubilization was specifically inhibited by the serine protease inhibitors aprotinin and pefabloc. Other protease inhibitors, including leupeptin and alpha 1-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 a 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 A beta.
Amyloid Animals