The resistance of electron-transport chain Fe–S clusters to oxidative damage during the reaction of peroxynitrite with mitochondrial complex II and rat-heart pericardium

The effects of peroxynitrite and nitric oxide on the iron–sulfur clusters in complex II (succinate dehydrogenase) isolated from bovine heart have been studied primarily by EPR spectroscopy and no measurable damage to the constitutive 2Fe–2S, 3Fe–4S, or 4Fe–4S clusters was observed. The enzyme can be repeatedly oxidized with a slight excess of peroxynitrite and then quantitatively re-reduced with succinate. When added in large excess, peroxynitrite reacted with at least one tyrosine in each subunit of complex II to form 3-nitrotyrosines, but activity was barely compromised. Examination of rat-heart pericardium subjected to conditions leading to peroxynitrite production showed a small inhibition of complex II (16%) and a greater inhibition of aconitase (77%). In addition, experiments performed with excesses of sodium citrate and sodium succinate on rat-heart pericardium indicated that the “g ∼ 2.01” EPR signal observed immediately following the beginning of conditions modeling oxidative/nitrosative stress, could be a consequence of both reversible oxidation of the constitutive 3Fe–4S cluster in complex II and degradation of the 4Fe–4S cluster in aconitase. However, the net signal envelope, which becomes apparent in less than 1 min following the start of oxidative/nitrosative conditions, is dominated by the component arising from complex II. Taking into account the findings of a previous study concerning complexes I and III (L.L. Pearce, A.J. Kanai, M.W. Epperly, J. Peterson, Nitrosative stress results in irreversible inhibition of purified mitochondrial complexes I and III without modification of cofactors, Nitric Oxide 13 (2005) 254–263) it is now apparent that, with the exception of the cofactor in aconitase, mammalian (mitochondrial) iron–sulfur clusters are surprisingly resistant to degradation stemming from oxidative/nitrosative stress.