Open AccessPurification and Characterization of the Cytochrome c Oxidase from Rhodopseudomonas sphaeroides
Author(s) -
GENNIS Robert B.,
CASEY Robert P.,
AZZI Angelo,
LUDWIG Bernd
Publication year - 1982
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1982.tb06667.x
Subject(s) - cytochrome c oxidase , rhodobacter sphaeroides , paracoccus denitrificans , biochemistry , heme a , oxidase test , cytochrome , respiratory chain , chemistry , vesicle , enzyme , biology , photosynthesis , membrane
When grown aerobically in the dark, Rhodopseudomonas sphaeroides develops a respiratory chain similar to that in mitochondria and the photosynthetic apparatus is suppressed. The aa 3 ‐type cytochrome c oxidase from Rps. sphaeroides has been purified in Triton X‐100 by affinity chromatography with Sepharose 4B coupled to yeast cytochrome c . The oxidase contains 14 nmol heme a/mg protein and is composed of three polypeptide subunits with relative molecular masses of 45000, 37000 and 35000. The enzyme is highly active in the presence of detergents, with a maximal velocity of 300 s −1 /mol oxidase using either yeast or horse‐heart cytochrome c . The Rps. sphaeroides oxidase is cross‐reactive with antibodies directed against the oxidases from Paracoccus denitrificans and Saccharomyces cerevisiae . A particularly close relationship is indicated in the case of P. denitrificans . The Rps. sphaeroides oxidase has been incorporated into phospholipid vesicles. The resulting oxidase in these vesicles demonstrates high enzymatic activity and a respiratory control ratio of 5. Using these vesicles, no evidence for proton extrusion accompanying cytochrome c oxidation was observed. The data suggest that the Rps. sphaeroides oxidase does not function as a proton pump.