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Inhibition of the erythrocyte (Ca2+ + Mg2+)-ATPase by nonheme iron
L Leclerc1, M Marden, C Poyart
1Institut National de la Santé et de la Recherche Médicale, Hôpital de Bicêtre, Le Kremlin-Bicêtre, France.
Insights
Nonheme iron inhibits erythrocyte calmodulin-stimulated (Ca2+ + Mg2+)-ATPase (CaM-ATPase) activity, likely via phospholipid peroxidation. This finding is crucial for understanding hemolytic anemias.
Area of Science:
- Biochemistry
- Cell Biology
- Hematology
Background:
- Congenital hemolytic anemias share oxidative processes.
- Erythrocyte calmodulin-stimulated (Ca2+ + Mg2+)-ATPase (CaM-ATPase) is inhibited in these anemias.
- Previous work showed hemin inhibits CaM-ATPase via thiol oxidation.
Purpose of the Study:
- To investigate the inhibitory effect of nonheme iron on CaM-ATPase activity.
- To elucidate the mechanism of nonheme iron-induced inhibition.
Main Methods:
- Enzyme activity assays of CaM-ATPase.
- Treatment with nonheme iron, hemin, butylated hydroxytoluene, and dithiothreitol.
- Analysis of inhibition mechanisms.
Main Results:
- Nonheme iron inhibits CaM-ATPase activity.
- Inhibition by nonheme iron is prevented by butylated hydroxytoluene, suggesting lipid peroxidation.
- Dithiothreitol does not restore activity, indicating the mechanism differs from hemin's thiol oxidation.
Conclusions:
- Nonheme iron inhibits erythrocyte CaM-ATPase, at least partly through peroxidation of membrane phospholipids.
- This mechanism contributes to CaM-ATPase dysfunction in hemolytic anemias.
Abstract:
The erythrocyte calmodulin-stimulated (Ca2+ + Mg2+)-ATPase (CaM-ATPase), an integral membrane protein, is inhibited in different types of congenital hemolytic anemias for which oxidative processes appear as a common feature. The oxidation of hemoglobin and its degradation lead to the accumulation of ferric heme (hemin) and nonheme iron in the red cell. We have shown previously that hemin inhibits the activity of the enzyme of normal erythrocyte (Leclerc et al. (1988) Biochim. Biophys. Acta, 946, 49-56) involving an oxidation of thiol groups. The present study demonstrates that nonheme iron also inhibits the CaM-ATPase activity. In contrast with hemin, the inhibition of the enzyme induced by the nonheme treatment is prevented by butylated hydroxytoluene, a protecting agent of unsaturated phospholipid peroxidations, while dithiothreitol, a reducing agent of protein disulfide bridges, does not restore the activity of the enzyme. We conclude that nonheme iron inhibits the enzyme at least in part, through the peroxidation of phospholipids of the membrane bilayer.