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. 2017 Feb 22;45(6):3323–3340. doi: 10.1093/nar/gkx127

Figure 6.

Figure 6.

Ppr10 and Mpa1 are colocalized in the mitochondrial matrix. (A) Ppr10 is localized in mitochondria. Mitochondria were isolated from cells expressing Ppr10-Myc and Mpa1-HA as described in Materials and Methods. Total cell extracts (T), mitochondria (Mt) and postmitochondrial supernatants (PMS) were analyzed by western blotting using anti-HA Ab to detect Mpa1-HA, anti-c-Myc Ab to detect Ppr10-Myc, anti-Trz2 and anti-Sla1 Abs. Trz2 is a mitochondrial protein; Sla1 is a nuclear marker. (B) Ppr10-Myc is protected against proteinase K in mitochondrial fractions. Isolated mitochondria were treated with proteinase K in the absence or presence of Triton X-100 as indicated. After precipitation with tricarboxylic acid, samples were analyzed by immunoblot analysis using anti-HA, anti-c-Myc, anti-FLAG and anti-HSP60 Abs. (C) Alkali treatment of mitochondria. Purified mitochondria were extracted with 0.1 M sodium carbonate, pH ∼11.5. After centrifugation, soluble proteins (Supernatant, S) and the membrane-bound proteins (Pellet, P) were analyzed by SDS-PAGE and western blotting for Mpa1-HA, Ppr10-Myc, the mitochondrial matrix protein Hsp60 and the mitochondrial inner membrane protein Cox2.