Figure 4.
The C terminus of Cox18p is protected from added protease by the inner membrane. (A) Mitochondria (MT) were isolated from a strain expressing a Cox18p-HA fusion protein (SCS61). The mitochondria were subjected to mitoplasting in the absence or presence of 100 μg/ml proteinase K (prot. K). Protein samples (75 μg per lane) were separated on a 12% SDS-PAGE gel and blotted. The blot was probed with anti-HA antibodies and then stripped and reprobed with anti-cytochrome b2 (cyt. b2) and anti-citrate synthase antibodies (cit. syn.). The C-terminal fragment of Cox18p-HA (fragment) produced in mitoplasts treated with protease is indicated. O.G., +1% octylglucoside. (B) A hydrophilicity plot of the Cox18p protein (Kyte and Doolittle, 1982). The line indicates the approximate length of the C-terminal fragment relative to the plot, taking into account the 3.5-kDa triple-HA tag.