FIGURE 3: Secretins are embedded in the mitochondrial OM and exposed to the cytosol. (A) Mitochondria isolated from wild type cells expressing the indicated secretins were either left untreated (Total) or subjected to alkaline extraction. The supernatant (S) and pellet (P) fractions were analysed by SDS–PAGE and immunodecoration with antibodies against the indicated proteins. Tom20, an integral OM protein; Aco1 and Hep1, soluble matrix protein. (B) Mitochondria isolated from wild type cells expressing the indicated secretins were treated with increasing amounts of proteinase K (PK). Samples were analysed by SDS–PAGE and immunodecoration with antibodies against PulD, HA-tag and the indicated mitochondrial proteins. Full-length PulD is indicated with an arrow whereas a proteolytic fragment of InvG with an asterisk.