The 12/15-lipoxygenase (12/15-LOX) enzyme introduces peroxyl groups, in a position-specific manner, into polyunsaturated fatty acids to form various kinds of bioactive lipid metabolites, including lipid-derived electrophiles (LDE). The resident peritoneal macrophage is the site of highest 12/15-LOX expression in the mouse. However, the role of the enzyme in the regulation of resident macrophages is not fully understood. Here, we describe a chemoproteomic method to identify the targets of enzymatically generated LDE. By treating mouse peritoneal macrophages with omega-alkynyl arachidonic acid (aAA), we identified a series of proteins adducted by LDE generated through a 12/15-LOX catalyzed reaction. Pathway analysis revealed a dramatic enrichment of proteins involved in energy metabolism and found that glycolytic flux and mitochondrial respiration were significantly affected by the expression of 12/15-LOX. Our findings thus highlight the utility of chemoproteomics using aAA for identifying intracellular targets of enzymatically generated LDE.
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