Figure 1.

Proteomic identification of SNX6 as a component of BACE1-harboring protein complex. A) Schematic representation of BACE1-TAP protein and purification strategy. TAP tag consists of the calmodulin binding domain, TEV protease cleavage site, and protein A, and was fused to the C terminus of BACE1. Inactive BACE1-TAP was generated by introducing a missense mutation (D92G) at the active site. B) Western blot analysis of SY5Y cells stably expressing either control vector (C) or BACE1-TAP (WT). Elevated C99 levels were observed in the stable SY5Y cells expressing BACE1-TAP. C99 was detected by 6E10 antibody, and asterisk denotes nonspecific bands. C) TAP process of BACE1-harboring complex prepared from formaldehyde (FA)-cross-linked SY5Y cell lysates. Samples from each purification step were analyzed by Western blotting using anti-BACE1 antibody. BACE1-TAP and BACE1 (D92G)-TAP cross-linked to higher molecular complexes.