Labeling of succinate-cytochrome c reductase with 125I. Accessibility of the peptides to the aqueous phases on the cytosolic and matrix sides of the mitochondrial membrane.
Lactoperoxidase-catalyzed radioiodination was used to study the arrangement of the component peptides of succinate-cytochrome c reductase with respect to the aqueous phases on each side of the mitochondrial inner membrane. Mitochondria depleted of their outer membrane and inside-out vesicles purified from submitochondrial particles by the lectin-affinity procedure (D'Souza, M. P., and Lindsay, J. G. (1981) Biochim. Biophys. Acta 640, 463-472) were iodinated using immobilized preparations of lactoperoxidase. The labeled membranes were solubilized in detergent and the succinate-cytochrome c reductase was purified by immunoprecipitation with specific IgG. Analysis of the radioiodine distribution after sodium dodecyl sulfate-polyacrylamide gel electrophoresis and comparison with peptide stain patterns show that bands 2 (64 kilodaltons), 6 (30 kilodaltons), 9 (15 kilodaltons), and 11 (less than 10 kilodaltons) are labeled from the cytoplasmic surface of the membrane. Bands 1 (72 kilodaltons), 4 (48 kilodaltons), and 8 (20 kilodaltons) appear to be labeled on the matrix side of the membrane, while bands 3 (52 kilodaltons), 5 (35 kilodaltons), 7 (25 kilodaltons), and 10 (11 kilodaltons) are labeled from both sides of the membrane. Tentative identification of the labeled bands suggests that band 1 is the large subunit of succinate dehydrogenase. Bands 3 and 4 represent proteins which have been referred to as core proteins I and II. Bands 5 and 6 are the proteins associated with cytochromes b and c1, respectively; band 7 is the Rieske iron-sulfur protein.
D000818:Animals; D010856:Columbidae; D003600:Cytosol / Q000201:enzymology; D007457:Iodine Radioisotopes; D008931:Mitochondria, Muscle / Q000201:enzymology*; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology*; D010088:Oxidoreductases / Q000378:metabolism*; D013053:Spectrophotometry; D013367:Submitochondrial Particles / Q000201:enzymology; D013384:Succinate Cytochrome c Oxidoreductase / Q000302:isolation & purification / Q000378:metabolism*
D007457:Iodine Radioisotopes; D010088:Oxidoreductases; D013384:Succinate Cytochrome c Oxidoreductase