EVIDENCE AGAINST A ROLE FOR THE KUNITZ DOMAIN IN AMYLOIDOGENIC AND SECRETORY PROCESSING OF THE AMYLOID PRECURSOR PROTEIN

The effect of the Kunitz proteinase inhibitor (KPI) on potential beta- amyloid precursor protein (beta PP)-processing activities from control and Alzheimer's disease (AD) brains was examined using fluorogenic su bstrates designed to mimic the secretory and amyloidogenic cleavages i n beta PP. In addition, the level of secretion of KPI-containing beta PP751 and KPI-lacking beta PP695 from transfected cells was examined t o assess the effect of the KPI on beta PP secretion. beta PP751 and be ta PP695, obtained from conditioned media of transfected cells, had no effect on proteinase activities against the secretory and amyloidogen ic substrates in extracts from control and AD brains. At similar conce ntrations beta PP751, but not beta PP695, completely inhibited the act ivity of trypsin against these substrates. Serine proteinase inhibitor s had only modest effects on activities from brain, whereas cysteine m odification completely inhibited them, indicating that these proteinas e activities were not of the serine type. Thus, the results do not sup port a role for the KPI in the secretion of beta PP or in the amyloido genic cleavage of beta PP. The amounts of beta PP695 and beta PP751 co llected from the media of transfected cells after 48 h of growth were similar, indicating an equal rate of secretion. This result suggests t hat the KPI domain in beta PP751 did not inhibit the secretory cleavag e in transfected cells.