FIGURE 3:
TMX1 selectively establishes mixed disulfides with membrane-bound BACE501. (A) MEFs were transfected with empty vector (EV), BACE501, BACE501Δ, HA-tagged TMX1, or TMX1C/A (lanes 1–5), BACE501 in combination with HA-tagged TMX1 or TMX1C/A (lanes 6 and 7), or BACE501Δ in combination with HA-tagged TMX1 or TMX1C/A (lanes 8 and 9). The [35S]methionine/cysteine-radiolabeled model substrates were immunoisolated from cell lysates with anti-BACE antibodies. The immunocomplexes were separated under nonreducing conditions. (B) Same as A, but immunocomplexes were separated under reducing conditions. (C) Same as A, for complexes immunoisolated with anti-HA. (D) Same as C, analysis under reducing conditions. pOx501, partially oxidized BACE501; pOx501Δ, partially oxidized BACE501Δ; fOx501Δ, fully oxidized BACE501Δ; fOx501, fully oxidized BACE501; Red501, reduced BACE501; Red501Δ, reduced BACE501Δ; DBC, disulfide-bonded complexes; MD, mixed disulfides; CNX, calnexin.