Studies on the Respiratory Chain-linked Reduced Nicotinamide Adenine Dinucleotide Dehydrogenase

The finding that ferredoxin and photosynthetic pyridine nucleotide reductase contain approximately equimolar amounts of “labile” sulfide and nonheme iron, and that both of these components are liberated in inorganic form with attendant inactivation upon acid treatment of these proteins (3-5), has stimulated considerable interest in the chemical nature and function of labile sulfide in oxidizing enzymes. This unusual type of sulfur has now been identified in several iron flavoproteins (xanthine and aldehyde oxidases and dihydroorotic dehydrogenase (6)), and very recently a hypothetical structure has been proposed for the linkages of iron and labile sulfide to the protein in bacterial ferredoxins (7). Although succinic dehydrogenase appears to have been the first enzyme in which the presence of labile sulfide and its close association with nonheme iron was recognized (8, 9), to date no information has been published on the concentration of this type of sulfur in succinic dehydrogenase or in the other iron flavoprotein components of the respiratory chain. This study was initiated in order to establish the quantitative relations of iron and labile sulfide in the reduced nicotinamide adenine dinucleotide and succinic dehydrogenases of the respiratory chain of mammalian heart and in the NADH-cytochrome c reductase fragments derived from the former enzyme by heat and acid-ethanol treatment. It is known that during thermal degradation of NADH dehydrogenase at moderate temperatures (lo), catalytic activity and the substrate-inducible electron paramagnetic resonance signal at g = 1.94 are lost at identical rates (11, 12). It was of considerable interest, therefore, to see whether a correlation existed between the liberation of labile sulfide from NADH dehydrogenase and the loss of catalytic activity and of the electron paramagnetic resonance signal during thermal degradation, since such a correlation would suggest the possible involvement of the sulfide-iron complex in the catalytic cycle of the enzyme. The results of this study are presented below.