FIGURE 2: Bacterial secretins are targeted within yeast cells mainly to mitochondria and form native-like complexes. (A) Whole cell lysate (WCL) and fractions corresponding to cytosol (C), light microsomes (ER) and mitochondria (M) were obtained from wild type yeast cells transformed with a plasmid encoding the indicated secretin. Samples were analysed by SDS–PAGE and immunodecoration with antibodies against PulD, HA tag, and the marker proteins Tom20, Tom70 or Fis1 for the mitochondrial fraction, Bmh1 or Hexokinase for the cytosol, and Erv2 or Sec61 for the microsomal/ER fraction. (B) Mitochondria were isolated from wild type yeast cells transformed with the indicated PulD variant. The isolated organelles were solubilized with 1% digitonin, 1% DDM, or 0.5% Triton X-100 and analysed on a 6-13% BN-PAGE followed by immunodecoration with antibodies against PulD. Short and long exposures are presented. High molecular weight oligomers are indicated by asterisks. (C) Microsomal (ER) or mitochondria (M) fraction isolated from wild type cells transformed with an empty plasmid (Ø) or a plasmid encoding HA-tagged InvG were lysed with 1% DDM and further analysed as described in part (B). Membranes of bacterial cells transformed with an empty plasmid (Ø) or a plasmid encoding FLAG-tagged InvG were treated and analysed in parallel in the same way. High molecular weight oligomers are indicated by asterisks.