Generation of the amyloid-beta peptide N terminus in Saccharomyces cerevisiae expressing human Alzheimer's amyloid-beta precursor protein

• Published in: The Journal of biological chemistry 1999-11, Vol.274 (48), p.33843-33846
• Main Authors: Greenfield, J P, Xu, H, Greengard, P, Gandy, S, Seeger, M
• Format: Article
• Language: English
• Subjects: Alzheimer Disease - metabolism; Amino Acid Sequence; Amyloid beta-Peptides - chemistry; Amyloid beta-Peptides - metabolism; Amyloid beta-Protein Precursor - analysis; Amyloid beta-Protein Precursor - genetics; Amyloid beta-Protein Precursor - metabolism; Amyloid Precursor Protein Secretases; Aspartic Acid Endopeptidases; b-Secretase; Electrophoresis, Polyacrylamide Gel; Endopeptidases - metabolism; Gene Expression; Humans; Molecular Weight; Peptide Fragments - chemistry; Peptide Fragments - metabolism; Precipitin Tests; Protein Processing, Post-Translational; Recombinant Fusion Proteins - analysis; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - chemistry; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism
• ISSN: 0021-9258
• DOI: 10.1074/jbc.274.48.33843

The Alzheimer's amyloid-beta precursor protein (betaAPP) is a type 1 membrane-spanning protein from which the Alzheimer's disease amyloid-beta peptide (Abeta) is proteolytically derived. To date, attempts to identify the enzymes responsible for Abeta generation have failed. Here we report the accumulation of Abeta-immunoreactive peptides in yeast expressing human betaAPP. Characterization of these peptides by metabolic labeling, immunoprecipitation with Abeta-specific antibodies, and N-terminal radiosequencing indicates that these peptides include the Abeta peptide at their N termini. The Abeta-like peptides generated in yeast were recovered predominantly as 8- and 12-14-kDa species. A 4-kDa species was recovered either when a protease-deficient strain was used to prevent breakdown or when the 8- and 12-14-kDa species were treated with disaggregating agents. The likely existence in yeast of enzymes generating the Abeta N terminus indicates that the molecular identification of yeast beta-secretase-like enzymes may be accomplished using genetic screens or empirical approaches based upon the sequenced genome of Saccharomyces cerevisiae.