Evidence for the inorganic nature of the cyanolyzable sulfur of molybdenum hydroxylases.
Activation of the desulfo forms of milk xanthine oxidase, chicken liver xanthine dehydrogenase, and aldehyde oxidase with S2- is greatly facilitated in the presence of reducing agents. Upon anaerobic incubation with 1 mM S2- and 1 mM dithionite, desulfo xanthine oxidase and chicken liver xanthine dehydrogenase prepared by cyanide treatment of active enzymes, are activated to the specific activity predicted by their molybdenum content. Routine preparations containing desulfo molecules are also similarly activated to the extent predicted. Cyanide-inactivated chicken liver xanthine dehydrogenase was reconstituted with 35S2- in the presence of dithionite. 85% of enzyme-bound radioactivity was shown to be in the form of cyanolyzable sulfur, by comparison of enzyme activity, bound radioactivity, and 35SCN- yields from exposure of labeled enzyme to cyanide. This radiolabeled enzyme allowed the determination of the following. 1) The cyanolyzable sulfur is largely removed from the polypeptide by incubation at 37 degrees C for one hour in 1% sodium dodecyl sulfate, pH 7, or for 15 min in 6 M guanidinium chloride, pH 6.2. 2) The cyanolyzable sulfur is "acid labile." [35S]Methylene blue is formed in the theoretical quantity from oxidized or substrate-reduced enzyme under the standard conditions for labile sulfur analysis by the methylene blue method. These data strongly support the conclusion that the cyanolyzable sulfur is a terminal sulfur ligand of the Mo atom, and is not part of an organic moiety.
D000332:Aerobiosis; D000444:Aldehyde Dehydrogenase; D000445:Aldehyde Oxidoreductases / Q000378:metabolism*; D000693:Anaerobiosis; D000818:Animals; D002417:Cattle; D002645:Chickens; D003486:Cyanides / Q000494:pharmacology*; D004578:Electron Spin Resonance Spectroscopy; D005260:Female; D007658:Ketone Oxidoreductases / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D008892:Milk / Q000201:enzymology; D008982:Molybdenum / Q000032:analysis*; D011817:Rabbits; D013455:Sulfur; D014968:Xanthine Dehydrogenase / Q000378:metabolism*; D014969:Xanthine Oxidase / Q000378:metabolism*
D003486:Cyanides; D013455:Sulfur; D008982:Molybdenum; D014968:Xanthine Dehydrogenase; D014969:Xanthine Oxidase; D000445:Aldehyde Oxidoreductases; D007658:Ketone Oxidoreductases; D000444:Aldehyde Dehydrogenase