Generation of amyloid beta protein from its precursor is sequence specific.
Cerebral deposition of amyloid beta protein (A beta) is an early and critical feature of Alzheimer's disease. Here we analyze the substrate requirements of proteases ("beta-secretases") that cleave the beta-amyloid precursor protein (beta APP) at the N-terminus of A beta (Asp-597 of beta APP695) in intact human cells. The cleavage requires a membrane-bound substrate but tolerates shifts in the distance of the hydrolyzed bond from the membrane. The major protease has a minimum recognition region of Val-594 to Ala-598; most substitutions in this sequence strongly decrease or eliminate A beta production. Only the Swedish familial Alzheimer's disease mutation (K595N/M596L) strongly increases A beta production. Moreover, in this mutant but not in the wild type, the entire cytoplasmic tail with its reinternalization signals can be deleted without affecting A beta N-terminal cleavage, consistent with the concept that cleavage of this mutant occurs in a different cellular compartment than that of wild-type molecules. Our results have important implications for current intensive approaches to develop assays for and identify enzymes with beta-secretase activity.
Citron
M
M
Department of Neurology, Harvard Medical School, Brigham and Women's Hospital, Boston, Massachusetts 02115.
Teplow
D B
DB
Selkoe
D J
DJ
D000544:Alzheimer Disease / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D053829:Amyloid Precursor Protein Secretases; D016229:Amyloid beta-Peptides / Q000096:biosynthesis* / Q000737:chemistry; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016282:Aspartic Acid Endopeptidases; D001483:Base Sequence; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D010450:Endopeptidases / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009447:Neuroblastoma; D009838:Oligodeoxyribonucleotides; D017354:Point Mutation; D011499:Protein Processing, Post-Translational; D013379:Substrate Specificity; D014162:Transfection; D014407:Tumor Cells, Cultured
D016229:Amyloid beta-Peptides; D016564:Amyloid beta-Protein Precursor; D009838:Oligodeoxyribonucleotides; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human