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Selective labeling of beef heart cytochrome oxidase subunit III with eosin‐5‐maleimide
Author(s) -
Müller Michele,
Azzi Angelo
Publication year - 1985
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(85)80664-5
Subject(s) - cytochrome c oxidase , maleimide , protein subunit , chemistry , eosin , biochemistry , medicine , enzyme , staining , pathology , gene , polymer chemistry
Cytochrome c oxidase has been isolated from beef heart mitochondria and labeled with the fluorochrome eosin‐5‐maleimide (EMA) after pretreatment with mersalyl. On SDS‐polyacrylamide gels, EMA fluorescence and absorption occurred at a single band corresponding to subunit III. Since only Cys 115 of the two cysteinyl residues of subunit III had been shown to be reactive towards water‐soluble SH‐reagents, it was concluded that this residue was the one labeled by EMA. The EMA/enzyme ratio was about 1. Gel filtration experiments have shown that upon treatment with dicyclohexylcarbodiimide, subunit III was loosened from the complex; this result suggests that the inhibitory effect of dicyclohexylcarbodiimide on the H + ‐translocation activity may be related to such a phenomenon.