Two transmembrane aspartates in presenilin-1 required for presenilin endoproteolysis and gamma-secretase activity

Accumulation of the amyloid-beta protein (A beta) in the cerebral cortex is an early and invariant event in the pathogenesis of Alzheimer's disease. T he final step in the generation of A beta from the beta-amyloid precursor p rotein is an apparently intramembranous proteolysis by the elusive gamma-se cretase(s)(1). The most common cause of familial Alzheimer's disease is mut ation of the genes encoding presenilins 1 and 2, which alters gamma-secreta se activity to increase the production of the highly amyloidogenic A beta, isoform(2). Moreover, deletion of presenilin-2 in mice greatly reduces gamm a-secretase activity(3), indicating that presenilin-1 mediates most of this proteolytic event. Here we report that mutation of either of two conserved transmembrane (TM) aspartate residues in presenilin-1, Asp 257 (in TM6) an d Asp 385 (in TM7), substantially reduces A beta production and increases t he amounts of the carboxy-terminal fragments of beta-amyloid precursor prot ein that are the substrates of gamma-secretase. We observed these effects i n three different cell lines as well as in cell-free microsomes. Either of the Asp-->Ala mutations also prevented the normal endoproteolysis of presen iIin-1 in the TM6 -->TM7 cytoplasmic loop. In a functional presenilin-1 var iant (carrying a deletion in exon 9) that is associated with familial Alzhe imer's disease and which does not require this cleavage(4), the Asp385-->Al a mutation still inhibited gamma-secretase activity. Our results indicate t hat the two transmembrane aspartate residues are critical for both presenil in-1 endoproteolysis and gamma-secretase activity, and suggest that preseni lin-1 is either a unique diaspartyl cofactor for gamma-secretase or is itse lf gamma-secretase, an autoactivated intramembranous aspartyl protease.