Recognition of mitochondrial protein precursor lacking arginine at position-2 by mitochondrial processing peptidase: Processing of bovine cytochrome P450(SCC) precursor

Mitochondrial processing peptidase (MPP) specifically cleaves off the N-ter minal presequence of the mitochondrial protein precursor. Previous studies demonstrated that Arg at position -2 from the cleavage site, which is found among many precursors, plays a critical role in recognition by MPP. We ana lyzed the structural elements of bovine cytochrome P450 side-chain cleavage enzyme precursor [pre-P450(SCC)], which has Ala at position -2, for recogn ition by MPP. Replacement of Ala position -2 of pre-P450(SCC) with Arg resu lted in an increase in the cleavage rate. Replacement with Gly caused a red uction in the cleavage rate and the appearance of an additional cleavage si te downstream of the authentic site. A pre-P450(SCC) mutant with Met at pos ition -2 retained cleavage efficiency equal to that of the wild type. These results indicate that -2 Ala of pre-P450(SCC) is recognized by MPP as a de terminant for precise cleavage, and that the amino acid at -2 is required t o have a straight methylene chain for interaction with the S-2 site. The pr eference for distal basic residues, a hydrophobic residue at +1, and hydrox yl residues at +2 and +3, was almost the same as those of the precursors wi th Arg at -2, indicating that the recognition mechanism of pre-P450(SCC) by MPP is essentially the same as that of the precursors with Arg at position -2.