FIGURE 4.

Effect of recombinant GSAP-16K in a membrane-based γ-secretase assay. a, recombinant GSAP-16K with a C-terminal His₆ tag was expressed in E. coli and purified on a nickel-nitrilotriacetic acid column. The protein was analyzed by SDS-PAGE under nonreducing (lane 1) and reducing (lane 2) conditions. b, membranes from SHSY5Y-SPA4CT cells were incubated for 4 h at 37 °C in the presence of BSA, GSAP-16K_His6, 0.5% DMSO, or 1 μm GSI and the level of Aβ40 generated was quantified by immunoassay. GSAP-16K_His6 was either untreated or pre-treated with 5 mm DTT for 30 min at 37 °C prior to the assay. Data are mean ± S.E. Statistical analysis was conducted using a one-way ANOVA with Dunnett's multiple comparison test (**, p < 0.001).