Formation of a cytochrome c-like species from horse apoprotein and hemin catalyzed by yeast mitochondrial cytochrome c synthetase.
Cytochrome c synthetase in yeast mitochondria catalyzes the formation of a yeast cytochrome c-like species from the apoprotein and hemin (Basile, G., DiBello, C., and Taniuchi, H. (1980) J. Biol. Chem. 255, 7181-7191). To test the specificity of this enzyme, 125I-labeled horse apocytochrome c was incubated with the yeast mitochondrial fraction in the presence of hemin, NADPH, and an ethanol extract of the postmitochondrial fraction. A radioactive 125I-labeled cytochrome c-like species was formed in yields of up to 26%. This 125I-labeled species is indistinguishable from horse cytochrome c by ion exchange chromatography (under the conditions which allow separation of horse and yeast cytochrome c), resistance in its reduced form to digestion by trypsin, resistance against autoxidation, reduction by cytochrome b2, and generation of the apoprotein after treatment with silver sulfate and dithiothreitol. With unlabeled horse apoprotein and [59Fe]hemin, the yield of a [59Fe-labeled horse cytochrome c-like species was up to 7% with respect to the apoprotein incubated. The yield of the 59Fe-labeled species was not altered by the addition of unlabeled FeCl3. Conversely, synthesis of the 59Fe-labeled species was not detectable after incubation of yeast mitochondria with unlabeled horse apoprotein, unlabeled hemin, and 59FeCl3. The formation of both 125I- and 59Fe-labeled cytochrome c-like species was sensitive to heat. Thus, we conclude that cytochrome c synthetase catalyzes direct bonding of heme (or hemin) to the apoprotein. Since the amino acid sequences of horse and yeast cytochromes c differ considerably, cytochrome c synthetase may recognize only a limited region(s) of the apoprotein.
D000818:Animals; D001059:Apoproteins / Q000378:metabolism*; D002474:Cell-Free System; D003574:Cytochrome c Group / Q000096:biosynthesis*; D006418:Heme / Q000378:metabolism*; D006736:Horses; D007501:Iron / Q000378:metabolism; D007700:Kinetics; D008190:Lyases; D008928:Mitochondria / Q000201:enzymology*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D014166:Transferases / Q000378:metabolism*
D001059:Apoproteins; D003574:Cytochrome c Group; D006418:Heme; D007501:Iron; D014166:Transferases; D008190:Lyases; C031581:cytochrome C synthetase