Figure 5.

Import mediated by the heme lyase targeting sequence occurs along the authentic heme lyase import pathway. The fusion proteins DCD 171–279 and DC₁D 101–169 were imported into isolated mitochondria. (A) The organelles were pretreated with 50 μg/ml trypsin (27). (B) ATP was hydrolyzed by treatment with 20 units/ml apyrase (ref. 27; import reactions contained 20 μM oligomycin and 5 μM carboxy-atractyloside). (C) The membrane potential was depleted by the addition of the uncoupling reagent carbonyl cyanide m-chlorophenylhydrazone (50 μM; CCCP; ref. 26). Under any of these conditions, import of presequence-containing preproteins into the matrix was strongly impaired (not shown). Further treatment of the samples and analysis of import were performed as described in Fig. 2. The result for only one of the heme lyase/DHFR fusion proteins is shown.