Crystal structures of mitochondrial processing peptidase reveal the mode for specific cleavage of import signal sequences

Background: Mitochondrial processing peptidase (MPP) is a metalloendopeptid ase that cleaves the N-terminal signal sequences of nuclear-encoded protein s targeted for transport from the cytosol to the mitochondria. Mitochondria l signal sequences vary in length and sequence, but each is cleaved at a si ngle specific site by MPP. The cleavage sites typically contain an arginine at position -2 (in the N-terminal portion) from the scissile peptide bond in addition to other distal basic residues, and an aromatic residue at posi tion +1. Mitochondrial import machinery recognizes amphiphilic helical conf ormations in signal sequences. However, it is unclear how MPP specifically recognizes diverse presequence substrates. Results: The crystal structures of recombinant yeast MPP and a cleavage-def icient mutant of MPP complexed with synthetic signal peptides have been det ermined. MPP is a heterodimer; its alpha and beta subunits are homologous t o the core II and core I proteins, respectively, of the ubiquinol-cytochrom e c oxidoreductase complex. Crystal structures of two different synthetic s ubstrate peptides cocrystallized with the mutant MPP each show the peptide bound in an extended conformation at the active site. Recognition sites for the arginine at position -2 and the +1 aromatic residue are observed. Conclusions: MPP bound two mitochondrial import presequence peptides in ext ended conformations in a large polar cavity. The presequence conformations differ from the amphiphilic helical conformation recognized by mitochondria l import components. Our findings suggest that the presequences adopt conte xt-dependent conformations through mitochondrial import and processing, hel ical for recognition by mitochondrial import machinery and extended for cle avage by the main processing component.