Enzymatic reduction of phospholipid and cholesterol hydroperoxides in artificial bilayers and lipoproteins.

Lipid hydroperoxides (LOOHs) in various lipid assemblies are shown to be efficiently reduced and deactivated by phospholipid hydroperoxide glutathione peroxidase (PHGPX), the second selenoperoxidase to be identified and characterized. Coupled spectrophotometric analyses in the presence of NADPH, glutathione (GSH), glutathione reductase and Triton X-100 indicated that photochemically generated LOOHs in small unilamellar liposomes are substrates for PHGPX, but not for the classical glutathione peroxidase (GPX). PHGPX was found to be reactive with cholesterol hydroperoxides as well as phospholipid hydroperoxides. Kinetic iodometric analyses during GSH/PHGPX treatment of photoperoxidized liposomes indicated a rapid decay of total LOOH to a residual level of 35-40%; addition of Triton X-100 allowed the reaction to go to completion. The non-reactive LOOHs in intact liposomes were shown to be inaccessible groups on the inner membrane face. In the presence of iron and ascorbate, photoperoxidized liposomes underwent a burst of thiobarbituric acid-detectable lipid peroxidation which could be inhibited by prior GSH/PHGPX treatment, but not by GSH/GPX treatment. Additional experiments indicated that hydroperoxides of phosphatidylcholine, cholesterol and cholesteryl esters in low-density lipoprotein are also good substrates for PHGPX. An important role of PHGPX in cellular detoxification of a wide variety of LOOHs in membranes and internalized lipoproteins is suggested from these findings.

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