Presenilin heterodimers apparently contain the active site of gamma-secretase, a 
polytopic aspartyl protease involved in the transmembrane processing of both the 
Notch receptor and the amyloid-beta precursor protein. Although critical to 
embryonic development and the pathogenesis of Alzheimer's disease, this protease 
is difficult to characterize, primarily because it is a multicomponent complex 
of integral membrane proteins. Here the functional gamma-secretase complex was 
isolated by using an immobilized active site-directed inhibitor of the protease. 
Presenilin heterodimers and nicastrin bound specifically to this inhibitor under 
conditions tightly correlating with protease activity, whereas several other 
presenilin-interacting proteins (beta-catenin, calsenilin, and 
presenilin-associated protein) did not bind. Moreover, anti-nicastrin antibodies 
immunoprecipitated gamma-secretase activity from detergent-solubilized 
microsomes. Unexpectedly, C83, the major endogenous amyloid-beta precursor 
protein substrate of gamma-secretase, was also quantitatively associated with 
the complex. These results provide direct biochemical evidence that nicastrin is 
a member of the active gamma-secretase complex, indicate that beta-catenin, 
calsenilin, and presenilin-associated protein are not required for gamma 
activity, and suggest an unprecedented mechanism of substrate-protease 
interaction.