The metabolism of beta-amyloid converting enzyme and beta-amyloid precursor protein processing.
Herein we investigated the processing of beta-secretase (BACE), implicated in Alzheimer's disease through processing of beta-amyloid precursor protein (betaAPP), into smaller metabolites. Four products of approximately 34, approximately 12, approximately 8, and approximately 5kDa were identified, none of which were generated autocatalytically. The approximately 34 and approximately 12kDa forms are held together by disulfide bridges. The approximately 34kDa form results from two cleavages: an N-terminal processing at RLPR(45) downward arrow by furin/PC5, and a C-terminal cleavage at SQDD(379) downward arrow by an unknown enzyme that also releases the C-terminal approximately 12kDa product. Microsequencing of the approximately 8 and approximately 5kDa fragments showed that they are the result of processing at VVFD(407) downward arrow and DMED(442) downward arrow, respectively. Mutagenesis of the identified cleavage sites revealed that the mutants D379A, D379L or D379E block the degradation of BACE into the approximately 12kDa product, confirming the importance of Asp(379). Notably, the D379E mutant results in higher betaAPP derived C99 levels. In contrast, D442A or D442E did not affect the production of the approximately 8 or approximately 5kDa products. The levels of the approximately 8 and approximately 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 betaAPP with BACE results in enhanced production of Abeta(11-40), the D407A mutant produces mostly Abeta(40).
Benjannet
Suzanne
S
Laboratory of Biochemical Neuroendocrinology, Clinical Research Institute of Montreal, 110 Pine Ave West, Montreal, Que., Canada.
Cromlish
James A
JA
Diallo
Karidia
K
Chrétien
Michel
M
Seidah
Nabil G
NG
D000595:Amino Acid Sequence; D053829:Amyloid Precursor Protein Secretases; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D016282:Aspartic Acid Endopeptidases; D001665:Binding Sites; D002460:Cell Line; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational
D016564:Amyloid beta-Protein Precursor; D007527:Isoenzymes; D010446:Peptide Fragments; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human; C509282:Bace1 protein, mouse