Positions of the heterodimeric toxin and its tetrameric derivative () 2 (which forms spontaneously under conditions of a nonreducing SDSPAGE) are indicated. (B) Immunoblot of secreted K28 toxin in cell-free culture supernatants of the temperature-sensitive yeast mutants sec71, sec72, sec62, sec63, and sec61 and their isogenic SEC wild-type equivalent after cultivation at the semirestrictive temperature of 30☌. Organellar marker proteins in the same subcellular fractions of the K28 killer strain MS300b were probed with anti-Kar2p (ER lumenal marker), anti-Sec61p (ER membrane marker), and anti-Pgk1p (cytosolic marker). Positions of the unprocessed pptox in the cytosol, the N-glycosylated ptox in the ER lumen, and the heterodimeric K28 toxin in toxin-treated (K28) and untreated (K28) cells are indicated. cerevisiae MS300b and its heat-cured nonkiller derivative probed with a polyclonal anti-subunit antibody. (A) SDSPAGE and Western blot analysis of the intracellular membrane fraction (M) and the cytosol (C) of the K28 superkiller mutant (ski2-2) S. Posttranslational pptox import into the ER and reinternalization of the mature toxin.

Prepros import