Human Cytoplasmic Aconitase (Iron Regulatory Protein 1) Is Converted into Its [3Fe-4S] Form by Hydrogen Peroxide in Vitro but Is Not Activated for Iron-responsive Element Binding

Abstract

Iron regulatory protein 1 (IRP1) regulates the synthesis of proteins involved in iron homeostasis by binding to iron-responsive elements (IREs) of messenger RNA. IRP1 is a cytoplasmic aconitase when it contains a [4Fe-4S] cluster and an RNA-binding protein after complete removal of the metal center by an unknown mechanism. Human IRP1, obtained as the pure recombinant [4Fe-4S] form, is an enzyme as efficient toward cis-aconitate as the homologous mitochondrial aconitase. The aconitase activity of IRP1 is rapidly lost by reaction with hydrogen peroxide as the [4Fe-4S] cluster is quantitatively converted into the [3Fe-4S] form with release of a single ferrous ion per molecule. The IRE binding capacity of IRP1 is not elicited with H(2)O(2). Ferrous sulfate (but not other more tightly coordinated ferrous ions, such as the complex with ethylenediamine tetraacetic acid) counteracts the inhibitory action of hydrogen peroxide on cytoplasmic aconitase, probably by replenishing iron at the active site. These results cast doubt on the ability of reactive oxygen species to directly increase IRP1 binding to IRE and support a signaling role for hydrogen peroxide in the posttranscriptional control of proteins involved in iron homeostasis in vivo.

Aconitase is the main functional target of aging in the citric acid cycle of kidney mitochondria from mice

“…For instance, exposure to superoxide and hydrogen peroxide ultimately causes the release of Fe-α from the [4Fe-4S] 2+ cluster, producing [3Fe-4S] 1+ and an inactivation of aconitase activity (Verniquet et al, 1991;Brazzolotto et al, 1999;Vasquez-Vivar et al, 2000;Bulteau et al, 2003). However, recent studies suggest that aconitase is likely to be inhibited by a post-translational modification since the amount of Fe-α removal does not reflect the magnitude of enzyme inactivation (Bulteau et al, 2003).…”

Non Transferrin Bound Iron: Nature, Manifestations and Analytical Approaches for Estimation

“…The fraction of iron which is inaccessible to Tf is the key cause of the NTBI existence at low Tf saturation [20]. Oxidative stress has also been documented as a cause as well as consequence of free iron, where the stable iron compound like iron sulphur proteins, ferritin and even haem release NTBI [34][35][36][37][38][39][40][41][42]. As shown in Table 1 free iron has been well documented as NTBI or other relevant subfractions in various iron overload conditions like hemochromatosis and thalassemia [11,20,[43][44][45].…”

Human iron regulatory protein 2 is easily cleaved in its specific domain: consequences for the haem binding properties of the protein

“…Absorption spectra, EPR spectra [28] and N-terminal sequences [29] were obtained as previously described.…”