Discovery of the Cysteine Protease Cathepsin B as a Drug Target for Alzheimer’s Disease - Proteinases as Drug Targets - page 150

Notably, the WT Lys (lysine) residue at the P2 position is a positively
charged hydrophilic residue, but the Swedish mutant Asn (asparagine) residue
is an uncharged amino acid. The charged and uncharged properties of Lys and
Asn, respectively, at the P2 position may be recognized by different proteases.
The P2 residue that interacts with the S2 subsite of proteases is known to be an
important determinant of substrate specificity for many proteases, such as the
cysteine cathepsins. 27 Based on the different charge property of the P2 residue
of the WT site compared to uncharged P2 residue of the Swedish mutant
b-secretase site, it is possible that different proteases may cleave the WT
b-secretase site compared to the Swedish mutant b-secretase site.
It is desirable to identify the protease(s) cleaving at the WT b-secretase site,
since it is expressed in 499% of the AD population. Therefore, consideration
of P1 to P3 residues for WT cleavage specificity can be designed as the peptide-
MCA substrate Z-Val-Lys-Met- k MCA. Cleavage at the peptide-like bond
liberates free, fluorescent AMC (aminomethylcoumarinamide).
6.3 Cathepsin B is Identified as a Candidate
b-Secretase in Secretory Vesicles for Production
of Ab Peptides
6.3.1 Regulated Secretory Vesicles Produce the Majority of
Secreted Ab
Accumulation of extracellular Ab results from neuronal production and
secretion of neurotoxic Ab that results in neurodegeneration of neurons in
brain regions (hippocampus and cortex) responsible for memory function
(Figure 6.1). Neurons possess the regulated secretory pathway that is utilized
for activity-dependent neurotransmitters, and the constitutive secretory path-
way for basal secretion. 14,30-32 It is important to understand the neurobiology
of Ab production with respect to the primary secretory pathway that provides
extracellular Ab. Data of in vivo secretion of Ab indicate its secretion primarily
from the regulated secretory pathway of neurons, summarized in this section
(Figure 6.1). The subcellular location of Ab production in regulated secretory
vesicles indicates that this organelle should be investigated for proteases that
generate Ab.
Studies in the field indicate that the majority of extracellular Ab in brain is
provided by activity-dependent secretion from neurons via the regulated
secretory pathway. Neurons undergo high-frequency, receptor-mediated, elec-
trical activity-dependent secretion of Ab. 15,33-35 Neurotransmitters also
undergo regulated secretion. 14,30-32 In parallel to the main regulated secretory
pathway for Ab secretion, a minor portion of Ab undergoes basal secretion
from the constitutive secretory pathway. 15 It is known from studies of peptide
neurotransmitter biosynthesis that different proteolytic enzymes are present in