EPR investigation of the active site of recombinant human 5-lipoxygenase: Inhibition by selenide

Lipoxygenases are a group of non-heme iron dioxygenases which catalyze the formation of lipid hydroperoxides from unsaturated fatty acids. 5-Lipoxygen ase (5LO) is of particular interest for formation of leukotrienes and lipox ins, implicated in inflammatory processes. In this study, electron paramagn etic resonance (EPR) spectroscopy was used to investigate the active site i ron of purified recombinant human 5-lipoxygenase (5LO), and to explore the action of selenide on 5LO. After oxidation by lipid hydroperoxides, 5LO exh ibited axial EPR spectra typified by a signal at g = 6.2. However, removal of the lipid hydroperoxides, their metabolites, and the solvent ethanol fro m the samples resulted in a shift to more rhombic EPR spectra (g = 5.17 and g = 9.0). Thus, many features of 5LO and soybean lipoxygenase-l EPR spectr a were similar, indicating similar flexible iron ligand arrangements in the se lipoxygenases. Selenide (1.5 muM) showed a strong inhibitory effect on t he enzyme activity of 5LO. In EPR, selenide abolished the signal at g = 6.2 , typical for enzymatically active 5LO. Lipid hydroperoxide added to seleni de-treated 5LO could not reinstate the signal at g = 6.2, indicating an irr eversible change of the coordination of the active site iron.