Figure 4
From: Binding of longer Aβ to transmembrane domain 1 of presenilin 1 impacts on Aβ42 generation
GSMs and P88L mutation alter interaction of TMD1 of PS1 with Aβ45. (A) Schematic representation of recombinant proteins used in this study. (B) GST and GST-fused recombinant proteins were purified by glutathione sepharose and/or Mono Q column. The purity of purified recombinant proteins was confirmed by silver staining. (C) GSM-1-BpB (4 nM) labeling experiment for GST-PS11-110 and TMD1 swapping mutant, GST-TM1mt1-110 (0.19 μg). Samples were preincubated with photoprobes on ice for 10 min, and then irradiated for 60 min. (D) Pull down assay of GST-fused recombinant proteins mixed with longer Aβ (Aβ43/45/46/48/49) (1 nM). After incubation with glutathione sepharose, all samples were washed and subjected to immunoblot. (E) The effects of GSM-1 (25 μM) or fenofibrate (100 μM) on the interaction of GST-PS11-110 with longer Aβ peptides. Recombinant proteins were preincubated with GSMs and then incubated with longer Aβ peptides. Protein complexes were pulled down by glutathione sepharose and analyzed by immunoblot. (F) The effect of P88L mutation on the binding of Aβ45 peptide to GST-PS11-110. (G) Binding of Aβ48 peptide to native PS1 protein expressed in mammalian cells. Densitometric analysis of relative levels of the bound Aβ48 was shown at the below (n = 3, mean ± SD, ***p <0.001 at Student’s t test).