Cytochrome c oxidase from bakers' yeast. IV. Immunological evidence for the participation of a mitochondrially synthesized subunit in enzymatic activity.
In order to study the role of the individual subunits of yeast cytochrome c oxidase, rabbit antisera were prepared against Subunit II (a mitochondrially made polypeptide) and Subunit VI (a cytoplasmically made polypeptide). Antisera were also obtained against a mixture of the two mitochondrially made subunits (I PLUS II) and against mixtures of the following cytoplasmically made subunits: (IV PLUS VI); (V PLUS VII); and (IV PLUS V PLUS VI PLUS VII). Neither anti-II serum nor anti-VI serum cross-reacted with any of the other six subunits of cytochrome c oxidase as judged by a sensitive ring test or by double diffusion in agarose gels. Anti-II serum inhibited the oxidation of ferrocytochrome c by purified yeast cytochrome c oxidase or by freshly isolated as well as sonically fragmented yeast mitochondria. Anti-(V, VII) serum and anti-(IV, V, VI, VII) serum were also strongly inhibitory. Anti-VI serum and anti-(IV, VI) serum inhibited only weakly. If purified cytochrome c oxidase was inhibited with a saturating amount of anti-VI serum, anti-II serum elicited a further increment of inhibition, as would be expected if the inhibitory effects of these two antisera involved different antigenic sites on the holoenzyme. Each of the antisera precipitated all seven cytochrome c oxidase subunits from crude mitochondrial extracts. However, anti-VI and, particularly, anti-II were much less effective precipitants than antisera against Subunits IV to VII or antisera against the holoenzyme. These data suggest that the oxidation of ferrocytochrome c by cytochrome c oxidase required both mitochondrially as well as cytoplasmically made subunits.
D000818:Animals; D003574:Cytochrome c Group; D003593:Cytoplasm / Q000201:enzymology; D003576:Electron Transport Complex IV / Q000096:biosynthesis / Q000276:immunology*; D005779:Immunodiffusion; D007700:Kinetics; D046911:Macromolecular Substances; D008928:Mitochondria / Q000201:enzymology*; D011817:Rabbits / Q000276:immunology; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D003574:Cytochrome c Group; D046911:Macromolecular Substances; D003576:Electron Transport Complex IV