EFFICIENT BUT ABERRANT CLEAVAGE OF MITOCHONDRIAL PRECURSOR PROTEINS BY THE CHLOROPLAST STROMAL PROCESSING PEPTIDASE

Cytosol-synthesised chloroplast and mitochondrial precursor proteins a re proteolytically processed after import by highly specific, metal-de pendent soluble enzymes: the stromal processing peptidase (SPP) and th e matrix processing peptidase (MPP), respectively. We have used in vit ro processing assays to compare the reaction specificities of highly p urified preparations of pea SPP and Neurospora crassa MPP, both of whi ch are unable to cleave a variety of 'foreign' proteins. We show that SPP can cleave all five mitochondrial precursor proteins tested, namel y cyclophilin, the beta subunit of the F-1-ATPase complex, the Rieske FeS protein, the alpha-MPP subunit and cytochrome b(2). In contrast, M PP is unable to cleave any chloroplast precursor proteins tested. Seve ral of the mitochondrial precursor proteins are cleaved more efficient ly by SPP than are many authentic chloroplast precursor proteins but, in each case, cleavage takes place at a site or sites which are N-term inal to the authentic MPP site; pre-cyclophilin is cleaved 5 residues upstream of the MPP site and the precursor of the beta subunit of the F-1-ATPase complex is cleaved at sites 5 and 12 residues upstream. We discuss the implications of these data for the SPP reaction mechanism.