Properties of the covalently bound flavin of chromatium cytochrome c-552 and its conversion to 8-carboxy-riboflavin

Previous work has shown that cytochrome c-552 from Chromatium contains a covalently bound flavin, which is not released by denaturation but is liberated by proteolysis or various chemical treatments and that the protein is probably attached at the 8α position of the FAD. In the present study unambiguous evidence was obtained for 8α substitution from (a) ESR hyperfine spectra of the flavin cation radical and (b) from identification of the flavin released by performic acid oxidation from cytochrome c-552 as riboflavin-8α-carboxylic acid. Various lines of evidence suggest that in the native enzyme a reduced sulfur may be linked to the 8α group of the flavin, similarly to monoamine oxidase, which contains cysteinyl 8α-FAD, although the conditions required for cleavage of the flavin from the two enzymes appear to be quite different.