Changes in γ-secretase activity and specificity caused by the introduction of consensus aspartyl protease active motif in Presenilin 1

Abstract

Presenilin (PS1 or PS2) is an essential component of the active γ-secretase complex that liberates the Aβ peptides from amyloid precursor protein (APP). PS1 is regarded as an atypical aspartyl protease harboring two essential aspartic acids in the context of the sequence D257LV and D385FI, respectively, rather than the typical DTG...DTG catalytic motif of classical aspartyl proteases. In the present studies, we introduced the sequence DTG in PS1 at and around the catalytic D257 and D385 residues to generate three PS1 mutants: D257TG, D385TG, and the double-mutant D257TG/D385TG. The effects of these changes on the γ-secretase activity in the presence or absence of γ-secretase inhibitors and modulators were investigated. The results showed that PS1 mutants having D385TG robustly enhanced Aβ 42 production compared to the wild type (wt), and were more sensitive than wt to inhibition by a classical aspartyl protease transition state mimic, and fenchylamine, a sulfonamide derivative. Unlike wt PS1 and some of its clinical mutants, all three PS1 artificial mutants decreased cleavage of Notch S3-site, suggesting that these artificial mutations may trigger conformational changes at the substrate docking and catalytic site that cause alteration of substrate specificity and inhibition pattern. Consistent with this notion, we have found that NSAID enzymatic inhibitors of COX, known modulators of the γ-secretase activity, cause PS1 mutants containing D385TG to produce higher levels of both Aβ 38 and Aβ 42 , but to reduce levels of Aβ 39 , showing a pattern of Aβ formation different from that observed with wild type PS1 and its clinical mutants. This study provides an important structural clue for the rational design of drugs to inhibit processing of APP at the γ-site without interfering with Notch processing.

Stepping closer to treating Alzheimer’s disease patients with BACE1 inhibitor drugs

“…AZD-3839 has 14 and >1000-fold greater selectivity over BACE2 and cathepsin D, respectively, and orally administrated AZD-3839 shows dose- and time-dependent lowering of plasma, brain, and CSF Aβ40, Aβ42, and sAPPβ levels in mouse (C57BL/6 WT mice), guinea pig, and non-human primate [ 42 ]. Interestingly, treatment with this compound showed a biphasic effect, which causes an initial small increase in Aβ levels followed by a subsequent significant reduction in Aβ, similar to our previous reports for γ-secretase inhibition [ 7 ]. It has also been reported that the IC 50 values to reduce Aβ40 in plasma were 64- and 48-fold lower than those in brains for mouse and guinea pig, respectively [ 42 ].…”

BACE and γ-Secretase Characterization and Their Sorting as Therapeutic Targets to Reduce Amyloidogenesis

“…7) without change in Notch at the S3 site cleavage (Fig. 8) that strongly points to conformation as a factor [419]. Catalysis resembles that of bacterial signal peptide peptidases (SPP) with a Gly-Asp 385 -X motif or polytopic type 4 prepilins [420][421][422] containing a conserved PAL motif [415].…”

A cellular model of amyloid precursor protein processing and amyloid-β peptide production

“…Differing by 2 amino acids at the carboxy-terminus, the choice of Aβ40 and Aβ42 isoform production is determined by γ-secretase cleavage site selection (Carter et al, 2008; Durkin et al, 1999). Notably, while Aβ40 accounts for approximately 90% of secreted Aβ and is the most abundant species recovered from cerebrospinal fluid, it is Aβ42 that is the predominant insoluble amyloid isoform deposited in the brain (Iwatsubo et al, 1994).…”