THE UBIQUINOL CYTOCHROME-C OXIDOREDUCTASE COMPLEX OF SPINACH LEAF MITOCHONDRIA IS INVOLVED IN BOTH RESPIRATION AND PROTEIN PROCESSING

Nuclear encoded mitochondrial precursor proteins are cleaved to mature size products by the general mitochondrial processing peptidase (MPP) . In contrast to non-plant tissues where MPP is located in the matrix, the general processing activity of potato tuber (storage tissue) mito chondria has been shown to constitute an integral part of the isolated cytochrome c reductase complex of the respiratory chain. Here we show isolation of MPP from photosynthetic tissue, spinach leaf mitochondri a, starting from the total membrane processing extract using extractio n with dodecyl-P-maltoside followed by FPLC anion-exchange and gel fil tration chromatography. The total spinach leaf MPP is found in the fra ctions containing the cytochrome c reductase complex and is shown to b e an integral part of the complex. No processing activity has been fou nd in any other fractions. The isolated cytochrome c reductase complex is shown to process three precursor proteins of different intramitoch ondrial localisation, the F,P subunit of ATP synthase (extrinsic membr ane protein on matrix side), the Rieske FeS protein (integral membrane protein facing intermembrane space) and the malate dehydrogenase (mat rix protein). The processing activity is totally inhibited by EDTA and orthophenanthroline. Our results together with the results in potato mitochondria show that integration of MPP into the cytochrome c reduct ase is a general phenomenon for plants. The complex consists of ten pr otein bands on SDS-PAGE of 61, 54, 52, 34, 32, 26, 15, 12, 11 and 10 k Da. The 61, 54 and 52 kDa bands correspond to Core proteins, the 32 kD a band to cytochrome b and the 26 kDa band to Rieske FeS protein as es timated by immunological methods. The three Core proteins are shown to be immunologically related to MPP from other sources, the Core 1 prot ein corresponding to beta-MPP and the Core 2 and Core 3 proteins corre sponding to alpha-MPP, which in comparison to MPP in potato mitochondr ia indicates species-dependent differences in the appearence of the pr ocessing components. Furthermore, the processing activity of the isola ted and membrane-bound spinach cytochrome c reductase complex is shown to be inhibited by antimycin A and myxothiazol, electron transfer inh ibitors of the complex. The inhibition of processing is, however, not correlated to the inhibition of electron transfer through the complex or to the redox state of the complex.