A cDNA encoding the smaller subunit (P52) of mitochondrial processing protease was isolated from a rat liver cDNA library, using cDNA fragment for yeast MAS1 as the probe. The deduced amino acid sequence is highly homologous to those of PEP from Neurospora crassa and MAS1 from Saccharomyces cerevisiae. After in vitro transcription and translation, the precursor peptide was imported into isolated rat liver mitochondria and processed to its mature form. © 1993 Academic Press. Inc.