Tim23 IMS domain mutants impair import into the matrix and sorting to the inner membrane differently. Mitochondria were isolated from wild-type control (WT) and tim23 mutant cells after cultivation in lactate medium at 30°C. Radiolabeled mitochondrial precursor proteins were incubated with those mitochondria at 30°C for indicated times. For quantification of the imported, protease-protected proteins, amounts of protease-resistant proteins in wild-type mitochondria after the longest incubation time were set to 100%. p, precursor form; i, processing-intermediate form; m, mature form.