Role of mitochondrial processing peptidase and AAA proteases in processing of the yeast acetohydroxyacid synthase precursor

We studied presequence processing of the mitochondrial‐matrix targeted acetohydroxyacid synthase (Ilv2). C‐terminal 3HA‐tagging altered the cleavage pattern from a single step to sequential two‐step cleavage, giving rise to two Ilv2‐3HA forms (A and B). Both cleavage events were dependent on the mitochondrial processing peptidase (MPP). We present evidence for the involvement of three AAA ATPases, m‐ and i‐AAA proteases, and Mcx1, in Ilv2‐3HA processing. Both, precursor to A‐form and A‐form to B‐form cleavage were strongly affected in a ∆yme1 mutant. These defects could be suppressed by overexpression of MPP, suggesting that MPP activity is limiting in the ∆yme1 mutant. Our data suggest that for some substrates AAA ATPases could play an active role in the translocation of matrix‐targeted proteins.