Human 15-lipoxygenase: induction by interleukin-4 and insights into positional specificity.
Arachidonate 15-lipoxygenase (15-lipoxygenase) is a lipid-peroxidizing enzyme associated with specific inflammatory cells seen in asthma and atherosclerosis. In atherosclerosis, 15-lipoxygenase is induced in the macrophages of human and rabbit lesions and has been implicated in foam cell formation. In human lung, 15-lipoxygenase is preferentially expressed in airway epithelial cells and eosinophils. Our studies have focused both on the regulation of expression and on the structure-function relationships of the enzyme. To determine factors that could regulate expression, peripheral blood monocytes were purified and cultured with combinations of 18 factors. Only interleukin-4 (60 pM) induced 15-lipoxygenase mRNA, protein and enzymatic activity. Interferon-gamma (100 pM) inhibited the interleukin-4 dependent induction of 15-lipoxygenase. Results with cultured human airway cells were similar. These data suggest that expression of 15-lipoxygenase is regulated by interleukin-4, and that 15-lipoxygenase is a potential downstream effector molecule for this potent cytokine. In parallel studies, we have investigated determinants of positional specificity using site-directed mutagenesis and bacterial expression of human 15-lipoxygenase. Hypotheses for mutagenesis were derived from an analysis of conserved differences among multiple lipoxygenase sequences. Switching four amino acids in 15-lipoxygenase to their counterparts in 12-lipoxygenase resulted in a variant enzyme that produced equal 12- and 15-lipoxygenation. Further analysis has identified two amino acids that completely control the positional specificity of 15-lipoxygenase. These data have led to a preliminary model of the enzyme's active site region.
Sigal
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Cardiovascular Research Institute, University of California, San Francisco 94143-0911.
Sloane
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Conrad
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DJ
D000595:Amino Acid Sequence; D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000737:chemistry / Q000235:genetics; D001093:Arachidonate 15-Lipoxygenase / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D002417:Cattle; D017124:Conserved Sequence; D004790:Enzyme Induction / Q000187:drug effects; D006801:Humans; D015847:Interleukin-4 / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009000:Monocytes / Q000201:enzymology; D011817:Rabbits; D051381:Rats; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D014132:Trachea / Q000201:enzymology
D015847:Interleukin-4; D001092:Arachidonate 12-Lipoxygenase; D001093:Arachidonate 15-Lipoxygenase