Figure 6.

COX3 activator Pet494p coimmuneprecipitates with the COX2 activator protein Pet111p. Purified mitochondria containing the indicated epitope-tagged proteins were treated with the cross-linker 2 mM DTME at 25°C for 1 h, followed by solubilization with 0.5% Triton X-100 at 4°C for 30 min. The clarified extracts (from 350 μg of mitochondria in each case) were incubated with the anti-Myc-agarose conjugate for 4 h at 4°C under gentle shaking conditions and immunoprecipitates were analyzed by Western blotting (see MATERIALS AND METHODS). Anti-HA-HRP and anti-Myc-HRP antibodies were used for probing the Western blots. Solubilized proteins from 20 μg of mitochondria were loaded in total extract lanes. Strains were CAB30 and SN33.