Use of diazido ethidium bromide as a specific probe for mitochondrial functions.
The diazido derivative of ethidium bromide has been synthesized as a potential photoaffinity label and shown to be at least as effective as a mitochondrial mutagen as the parent compound, with a similar mode of action. Exposure of mitochondria of Saccharomyces cerevisiae to the compound, followed by ultraviolet-irradiation, which converts it to the highly reactive dinitrene, results in its specific binding to a single component which has been tentatively identified as the smallest polypeptide (subunit 9) of the membrane-bound ATPase. An analogus reaction is also obtained with the soluble, oligomycin-sensitive ATPase complex but not with the F1-ATPase itself. The reaction with the ATPase complex can also be monitored by fluorescence enhancement and by this attribute, as well as by other criteria, diazido-ethidium bromide, ethidium bromide itself, euflavine, N,N'-dicyclohexylcarbodiimide, 2,4-dinitrophenol, and 2-azido-4-nitrophenol all appear to compete for the same, lipophilic, binding site. A mitochondrial mutation (73/1) (see Flury, U., Feldman, F., and Mahler, H.R. (1974) J. Biol. Chem. 249, 6630-6637) produces a photoaffinity product with an altered electrophoretic mobility and molecular weight.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D001386:Azides / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004996:Ethidium / Q000031:analogs & derivatives / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism; D007700:Kinetics; D008055:Lipids / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009840:Oligomycins / Q000494:pharmacology; D011955:Receptors, Drug / Q000187:drug effects; D012441:Saccharomyces cerevisiae / Q000378:metabolism*
D001386:Azides; D005656:Fungal Proteins; D008055:Lipids; D009840:Oligomycins; D011955:Receptors, Drug; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D004024:Dicyclohexylcarbodiimide; D000251:Adenosine Triphosphatases; D004996:Ethidium