REDOX PROPERTIES OF IRON IN THE BINDING SITE(S) OF F(1)ATPASE FROM MAMMALIAN MITOCHONDRIA AND THERMOPHILIC BACTERIUM PS3 - A COMPARATIVE-STUDY

Iron ions in the two iron centers of beef heart mitochondrial F(1)ATPa se, which we have been recently characterized (FEBS Letters 1996, 379, 231-235), exhibit different redox properties. In fact, the ATP-depend ent site is able to maintain iron in the redox state of Fe(II) even in the absence of reducing agents, whereas in the nucleotide-independent site iron is oxidized to Fe(III) upon removal of the reductant. Fe(II I) ions in the two sites display different reactivity towards H2O2, be cause only Fe(III) bound in the nucleotide-independent site rapidly re acts with H2O2 thus mediating a 30% enzyme inactivation. Thermophilic bacterium PS3 bears one Fe(III) binding site, which takes up Fe(III) e ither in the absence or presence of nucleotides and is unable to maint ain iron in the redox state of Fe(II) in the absence of ascorbate. Fe( III) bound in thermophilic F(1)ATPase in a molar ratio 1:1 rapidly rea cts with H2O2 mediating a 30% enzyme inactivation. These results suppo rt the presence in mitochondrial and thermophilic F(1)ATPase of a cons erved site involved in iron binding and in oxidative inactivation, in which iron exhibits similar redox properties. On the other hand, at va riance with thermophilic F(1)ATPase, the mitochondrial enzyme has the possibility of maintaining one equivalent of Fe(II) in its peculiar AT P-dependent site, besides one equivalent of Fe(III) in the conserved n ucleotide-independent site. In this case mitochondrial F(1)ATPase unde rgoes a higher inactivation (75%) upon exposure to H2O2 Under all cond itions the inactivation is significantly prevented by PEN and DMSO but not by Cu, Zn superoxide dismutase, thus suggesting the formation of OK radicals as mediators of the oxidative damage. No dityrosines, carb onyls or oxidized thiols are formed. Ln addition, in any cases no prot ein fragmentation or aggregation is observed upon the treatment with H 2O2.