Amyloid beta-precursor protein (APP) proteolytic products C83 and C99 are substrates for gamma-secretase as well as products, respectively, of alpha- or beta-secretase. In contrast to APP, C83 and C99 were derivatized by a water soluble biotinylation reagent to a much greater extent at 18 degrees C than at 0 degrees C in CHO cells expressing the Swedish mutant form of APP750. Intracellular C99 and C83 cycle to the cell surface when maintained in buffered saline at 18 degrees C thus identifying proteins derivatized at 18 degrees C as residing in recycling compartments. More than 80% of C99 and C83 biotinylated at 18 degrees C is associated with detergent resistant membrane (DRM). There thus appears to be no differential distribution of alpha- or beta-secretase products into the DRM fraction that would be expected if localization to DRM determines alternative secretase pathways. gamma-Secretase inhibitors increased the fraction of C99 but not C83 in the 18 degrees C pool by >50% and doubled the half-life of C99 in that compartment, showing that a substantial amount of C99 is proteolyzed by gamma-secretase in a compartment rich in recycling proteins. The temporal appearance of APP on the cell surface preceded that of C99 in the recycling compartment, further supporting the cleavage of APP in recycling endosomes. (C) 2010 Elsevier Inc. All rights reserved.