PT - JOURNAL ARTICLE AU - Mako Takami AU - Yu Nagashima AU - Yoshihisa Sano AU - Seiko Ishihara AU - Maho Morishima-Kawashima AU - Satoru Funamoto AU - Yasuo Ihara TI - γ-Secretase: Successive Tripeptide and Tetrapeptide Release from the Transmembrane Domain of β-Carboxyl Terminal Fragment AID - 10.1523/JNEUROSCI.2362-09.2009 DP - 2009 Oct 14 TA - The Journal of Neuroscience PG - 13042--13052 VI - 29 IP - 41 4099 - http://www.jneurosci.org/content/29/41/13042.short 4100 - http://www.jneurosci.org/content/29/41/13042.full SO - J. Neurosci.2009 Oct 14; 29 AB - Amyloid β protein (Aβ), a pathogenic molecule associated with Alzheimer's disease, is produced by γ-secretase, which cleaves the β-carboxyl terminal fragment (βCTF) of β-amyloid precursor protein in the middle of its transmembrane domain. How the cleavage proceeds within the membrane has long been enigmatic. We hypothesized previously that βCTF is cleaved first at the membrane–cytoplasm boundary, producing two long Aβs, Aβ48 and Aβ49, which are processed further by releasing three residues at each step to produce Aβ42 and Aβ40, respectively. To test this hypothesis, we used liquid chromatography tandem mass spectrometry (LC-MS/MS) to quantify the specific tripeptides that are postulated to be released. Using CHAPSO (3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxyl-1-propanesulfonate)-reconstituted γ-secretase system, we confirmed that Aβ49 is converted to Aβ43/40 by successively releasing two or three tripeptides and that Aβ48 is converted to Aβ42/38 by successively releasing two tripeptides or these plus an additional tetrapeptide. Most unexpectedly, LC-MS/MS quantification revealed an induction period, 3–4 min, in the generation of peptides. When extrapolated, each time line for each tripeptide appears to intercept the same point on the x-axis. According to numerical simulation based on the successive reaction kinetics, the induction period exists. These results strongly suggest that Aβ is generated through the stepwise processing of βCTF by γ-secretase.