The metabolism of β-amyloid converting enzyme and β-amyloid precursor protein processing

• Published in: Biochemical and biophysical research communications 2004-12, Vol.325 (1), p.235-242
• Main Authors: Benjannet, Suzanne, Cromlish, James A., Diallo, Karidia, Chrétien, Michel, Seidah, Nabil G.
• Format: Article
• Language: English
• Subjects: Alzheimer's disease; Amino Acid Sequence; Amyloid beta-Protein Precursor - chemistry; Amyloid beta-Protein Precursor - genetics; Amyloid beta-Protein Precursor - metabolism; Amyloid Precursor Protein Secretases; Animals; Asp cleavage site; Aspartic Acid Endopeptidases; Aβ formation; Binding Sites; Cell Line; Endopeptidases - genetics; Endopeptidases - metabolism; ER/Golgi intermediate compartment; Humans; Isoenzymes - genetics; Isoenzymes - metabolism; Metabolism; Mice; Microsequencing; Molecular Weight; Mutagenesis; Mutagenesis, Site-Directed; Peptide Fragments - genetics; Peptide Fragments - metabolism; Protein Processing, Post-Translational; β-Secretase; βAPP processing
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2004.10.019

Herein we investigated the processing of β-secretase (BACE), implicated in Alzheimer's disease through processing of β-amyloid precursor protein (βAPP), into smaller metabolites. Four products of ∼34, ∼12, ∼8, and ∼5kDa were identified, none of which were generated autocatalytically. The ∼34 and ∼12kDa forms are held together by disulfide bridges. The ∼34kDa form results from two cleavages: an N-terminal processing at RLPR45↓ by furin/PC5, and a C-terminal cleavage at SQDD379↓ by an unknown enzyme that also releases the C-terminal ∼12kDa product. Microsequencing of the ∼8 and ∼5kDa fragments showed that they are the result of processing at VVFD407↓ and DMED442↓, respectively. Mutagenesis of the identified cleavage sites revealed that the mutants D379A, D379L or D379E block the degradation of BACE into the ∼12kDa product, confirming the importance of Asp379. Notably, the D379E mutant results in higher βAPP derived C99 levels. In contrast, D442A or D442E did not affect the production of the ∼8 or ∼5kDa products. The levels of the ∼8 and ∼5kDa products are significantly lower in the mutant D407A but less so D407E, likely due to the low efficacy of ER exit of the D407A mutant. Indeed, while co-expression of βAPP with BACE results in enhanced production of Aβ11–40, the D407A mutant produces mostly Aβ40.