Glu 191 and Asp 195 in Rat Mitochondrial Processing Peptidase β Subunit Are Involved in Effective Cleavage of Precursor Protein through Interaction with the Proximal Arginine

Mitochondrial processing peptidase (MPP), consisting of α and β subunits, recognizes a large variety of N-terminal extension peptides of mitochondrial precursor proteins, and generally cleaves a single site of the peptide including arginine at the −2 position (P 2 ). We obtained evidence that Glu 191 and Asp 195 of rat β subunit interact with P 2 arginine of precursor protein through ionic and hydrogen bonds, respectively, using recombinant MPP. Mutation to alanines at Glu 191 and Asp 195 reduced processing activity toward precursors with P 2 arginine, but resulted in no loss of activity toward P 2 alanine precursors. Charge-complementary mutation demonstrated that MPP variants with β Arg 191 exhibited compensatory processing activity for the precursor with acidic residue at the P 2 position. Thus, Glu 191 and Asp 195 are substrate-binding sites required for cleavage of extension peptides through interaction with P 2 arginine.