Distinct pharmacological effects of inhibitors of signal peptide peptidase and gamma-secretase.

Signal peptide peptidase (SPP) and gamma-secretase are intramembrane aspartyl proteases that bear similar active site motifs but with opposite membrane topologies. Both proteases are inhibited by the same aspartyl protease transition-state analogue inhibitors, further evidence that these two enzymes have the same basic cleavage mechanism. Here we report that helical peptide inhibitors designed to mimic SPP substrates and interact with the SPP initial substrate-binding site (the "docking site") inhibit both SPP and gamma-secretase, but with submicromolar potency for SPP. SPP was labeled by helical peptide and transition-state analogue affinity probes but at distinct sites. Nonsteroidal anti-inflammatory drugs, which shift the site of proteolysis by SPP and gamma-secretase, did not affect the labeling of SPP or gamma-secretase by the helical peptide or transition-state analogue probes. On the other hand, another class of previously reported gamma-secretase modulators, naphthyl ketones, inhibited SPP activity as well as selective proteolysis by gamma-secretase. These naphthyl ketones significantly disrupted labeling of SPP by the helical peptide probe but did not block labeling of SPP by the transition-state analogue probe. With respect to gamma-secretase, the naphthyl ketone modulators allowed labeling by the transition-state analogue probe but not the helical peptide probe. Thus, the naphthyl ketones appear to alter the docking sites of both SPP and gamma-secretase. These results indicate that pharmacological effects of the four different classes of inhibitors (transition-state analogues, helical peptides, nonsteroidal anti-inflammatory drugs, and naphthyl ketones) are distinct from each other, and they reveal similarities and differences with how they affect SPP and gamma-secretase.