title: Cleavage of precursors by the mitochondrial processing peptidase requires a compatible mature protein or an intermediate octapeptide.
abstract: Many precursors of mitochondrial proteins are processed in two successive steps by independent matrix peptidases (MPP and MIP), whereas others are cleaved in a single step by MPP alone. To explain this dichotomy, we have constructed deletions of all or part of the octapeptide characteristic of a twice cleaved precursor (human ornithine transcarbamylase [pOTC]), have exchanged leader peptide sequences between once-cleaved (human methylmalonyl-CoA mutase [pMUT]; yeast F1ATPase beta-subunit [pF1 beta]) and twice-cleaved (pOTC; rat malate dehydrogenase (pMDH); Neurospora ubiquinol-cytochrome c reductase iron-sulfur subunit [pFe/S]) precursors, and have incubated these proteins with purified MPP and MIP. When the octapeptide of pOTC was deleted, or when the entire leader peptide of a once-cleaved precursor (pMUT or pF1 beta) was joined to the mature amino terminus of a twice-cleaved precursor (pOTC or pFe/S), no cleavage was produced by either protease. Cleavage of these constructs by MPP was restored by re-inserting as few as two amino-terminal residues of the octapeptide or of the mature amino terminus of a once-cleaved precursor. We conclude that the mature amino terminus of a twice-cleaved precursor is structurally incompatible with cleavage by MPP; such proteins have evolved octapeptides cleaved by MIP to overcome this incompatibility.
authors.affiliation: Yale University School of Medicine, Department of Human Genetics, New Haven, Connecticut 06510.
authors.forename: G
authors.initials: G
authors.lastname: Isaya
authors.forename: F
authors.initials: F
authors.lastname: Kalousek
authors.forename: W A
authors.initials: WA
authors.lastname: Fenton
authors.forename: L E
authors.initials: LE
authors.lastname: Rosenberg
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002451:Cell Compartmentation; D004252:DNA Mutational Analysis; D010450:Endopeptidases / Q000378:metabolism*; D066298:In Vitro Techniques; D007506:Iron-Sulfur Proteins / Q000378:metabolism; D008291:Malate Dehydrogenase / Q000378:metabolism; D008666:Metalloendopeptidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D008765:Methylmalonyl-CoA Mutase / Q000378:metabolism; D008930:Mitochondria, Liver / Q000201:enzymology / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009954:Ornithine Carbamoyltransferase / Q000378:metabolism; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013329:Structure-Activity Relationship; D000096122:Mitochondrial Processing Peptidase
chemical_list: D010450:Endopeptidases; D007506:Iron-Sulfur Proteins; D008291:Malate Dehydrogenase; D008666:Metalloendopeptidases; D008765:Methylmalonyl-CoA Mutase; D009954:Ornithine Carbamoyltransferase; D011498:Protein Precursors; D011994:Recombinant Proteins; D000096122:Mitochondrial Processing Peptidase; C068038:mitochondrial intermediate peptidase