Unique enzyme specificity of three human phospholipases A2 toward phospholipids containing sn‐2 omega‐3 and omega‐6 fatty acids

Abstract

Phospholipase A2 (PLA2) constitutes a superfamily of enzymes that hydrolyzes the sn‐2 acyl‐chain of membrane phospholipids producing lyso‐phospholipids and free fatty acids. The products of PLA2 have various functions, especially omega‐3 and omega‐6 fatty acids which are precursor of various eicosanoids and related metabolites involved in numerous pro‐inflammatory and pro‐resolving cellular responses. The PLA2 superfamily is composed of six different types of enzymes, each with distinct structural features. Among them, the cytosolic PLA2 (cPLA2), calcium‐independent PLA2 (iPLA2), and secreted PLA2 (sPLA2) are well studied. cPLA2 is the main provider of free arachidonic acid (AA), an omega‐6 fatty acid involved in the inflammatory cascade. Several knockout mice studies demonstrated the contribution of iPLA2 to the metabolism of the omega‐3 fatty acid docosahexaenoic acid (DHA) in brain (J. Lipid Res. 2010, 51, 3166–3173). sPLA2 is known for its antibacterial action and involvement in atherosclerosis. Previous studies in our laboratory determined the substrate specificity of all three PLA2s toward a variety of membrane phospholipids (J. Am. Chem. Society 2018 140, 3285–3291). However, the specificity of these enzymes toward phospholipids containing omega‐3 and omega‐6 fatty acids at the sn‐2 position has been unclear. In this study, we have employed a lipidomics‐based LC‐MS/MS assay, which measures the lysophospholipid product to measure the activity of PLA2, and to investigate the sn‐2 acyl‐chain specificity in various phospholipids. We have also developed a lipidomics‐based GC‐MS assay which allow us to measure free fatty acids in mixtures of phospholipid substrates. Although cPLA2 showed relatively high activity toward the omega‐3 eicosapentaenoic acid (EPA), it prefers AA. iPLA2 showed a distinct preference for EPA over AA and a much lesser activity toward phospholipids containing DHA. Unlike the other two PLA2s, sPLA2 showed a remarkable preference for DHA among the omega‐3 and omega‐6 fatty acids. Importantly, similar sn‐2 acyl‐chain specificity was obtained in mixtures of phospholipids. This study investigated the preference of three human PLA2 enzymes for the sn‐2 omega‐3 and omega‐6 fatty acids for the first time. Our findings will help us to understand the difference of function of PLA2 enzymes at the cellular level and their implications in inflammatory diseases.Support or Funding InformationSupported by NIH grant GM20501This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.