These data suggest that the lack of catalytic activity is the major reason for the embryonic lethality of Gpx4(-/-) mice and that systemic inactivation of the Alox15 gene does not rescue homozygous knock-in mice expressing catalytically silent Gpx4.

The oxidation of low density lipoprotein (LDL) by mammalian 15-lipoxygenases (15-LOX) was implicated in early atherogenesis. We investigated the molecular mechanism of 15-LOX/LDL interaction and found that during short term incubations, LDL cholesterol esters are oxygenated preferentially by the enzyme. Even when the LDL particle was loaded with free linoleic acid, cholesteryl linoleate constituted the major LOX substrate. In contrast, only small amounts of free oxygenated fatty acid isomers were detected, and re-esterification of oxidized fatty acids into the LDL ester lipid fraction was ruled out. When LDL was depleted from ␣-tocopherol, specific oxygenation of the cholesterol esters was not prevented, and the product pattern was not altered. Similar results were obtained at low (LDL/LOX ratio of 1:1) and high LOX loading (LDL/LOX ratio of 1:10) of the LDL particle. During long term incubations (up to 24 h), a less specific product pattern was observed. However, when the hydroperoxy lipids formed by the 15-LOX were immediately reduced by the phospholipid hydroperoxide glutathione peroxidase, when the reaction was carried out with vitamin E-depleted LDL, or when the assay sample was diluted, the specific pattern of oxygenation products was retained over a long period of time.These data suggest that mammalian 15-LOX preferentially oxidize LDL cholesterol esters, forming a specific pattern of oxygenation products. During long term incubations, free radical-mediated secondary reactions, which lead to a more unspecific product pattern, may become increasingly important. These secondary reactions appear to be suppressed when the hydroperoxy lipids formed are immediately reduced, when ␣-tocopherol-depleted LDL was used, or when the incubation sample was diluted. It may be concluded that 15-LOXinitiated LDL oxidation constitutes a dual-type oxygenase reaction with an initial enzymatic and a subsequent nonenzymatic phase. The biological relevance of this dual-type reaction for atherogenesis will be discussed. (15). (vi) A 15-LOX inhibitor that apparently lacks major antioxidative properties prevented lipid deposition in the aorta of cholesterolfed rabbits (16). These data are consistent with a proatherogenic activity of the enzyme. On the other hand, transgenic rabbits that overexpress the 15-LOX specifically in monocyte/ macrophages (17) develop significantly less atherosclerotic lesions when fed a cholesterol-rich diet or when cross-bred with LDL receptor-deficient Watanabe rabbits (18), suggesting an anti-atherogenic action of the enzyme.For a better understanding of the processes involved in LDL oxidation in vivo, the in vitro interaction of 15-LOXs with human LDL has been studied. Sparrow et al. (6) have shown that the 15-LOX from soybeans is capable of oxidizing LDL in the presence of phospholipase A 2 . Later on, we reported that the native rabbit (19) and the recombinant human 15-LOX (20) specifically oxidize LDL ester lipids in the absence of lipidhydrolyzing enzymes. Comparison of various LOX isoenzymes ind...

The lipoxygenase from reticulocytes oxygenates 1 SLs-HETE to 8-hydroperoxy-15-hydroxy-5,9,11,13-eicosatetraenoic acid and 5-hydroperoxy-15-hydroxy-6,8,11,13_eicosatetraenoic acid only in the presence of catalytic concentrations of monohydroperoxy fatty acids. During this reaction the hydroperoxy fatty acids are converted to more polar products including hydroxy fatty acids. From kinetic measurements of 15Ls-HETE oxygenation it was calculated that 1 mol monohydroperoxy fatty acid is consumed during the oxygenation of about 9 mol 1 SLs-HETE.Reticulocyte Lipoxygenase

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