FIGURE 3.

Import of CYP proteins into yeast mitochondria with deleted TOM subunits. A–E, mitochondria were isolated from wild-type, tom70Δ, tom20Δ, or tom22Δ yeast strains. ³⁵S-Labeled proteins (CYP+33/CYP1A1 (A), CYP2B1 (B), CYP+5/1A1 (C), CYP2E1 (D), and CYP27A1 (E)) were incubated with mitochondria from either wild-type or TOM deletion strains as indicated. Following import, mitochondria were treated with trypsin, re-isolated, and analyzed by SDS-PAGE and fluorography. The radioactive proteins bound to mitochondria and the protease-protected fractions were quantified for calculations of the % import (values presented at the bottom of lanes). F, ³⁵S-labeled CYP+331A1 and CYP2E1 were imported into tom22Δ mitochondria as described for A–E and resolved on a higher resolution gradient gel. G, trypsin- and digitonin (75 μg/mg protein)-treated fractions were also analyzed for marked proteins by immunoblot analysis using indicated antibodies. A and D, ← indicates the shorter protected fragment.