Mitochondrial proteinases of the yeast Saccharomyces cerevisiae: electrophoretic detection, substrate specificity and sensitivity to inhibitors

The proteins of submitochondrial particles solubilized with 0.1% Triton X-100 were separated by polyacrylamide gel electrophoresis. Hydrolysis of several proteinase substrates was registered directly in the gel after completion of electrophoresis. According to the data obtained the inner mitochondrial membrane contains one or two enzymes which catalyze hydrolysis of cytochrome c as well as one or two enzymes splitting synthetic substrate of trypsin-like proteinases, e. g. N-alpha-benzoyl-L-arginine-p-nitroanilide (BAPA) and N-alpha-benzoyl-L-arginine-beta-naphthylamide (BANA). Submitochondrial particles were shown to catalyze hydrolysis of 3H-labelled cytochrome c. This activity is suppressed by the same inhibitors as the hydrolysis of mitochondrial translation products, i. e. phenyl-methylsulfonylfluoride, p-chloromercuribenzosulfonate, leupeptin and antipain. Presumably these two processes are catalyzed by the same enzyme localized in the inner mitochondrial membrane. Physiological functions of BAPA- and BANA-hydrolyzing enzyme(s) are still unclear.