Omega-3 versus Omega-6 fatty acid availability is controlled by hydrophobic site geometries of phospholipase A2s
Daiki Hayashi, Varnavas D. Mouchlis, Edward A. Dennis
Abstract
Human phospholipase A 2 s (PLA 2 ) constitute a superfamily of enzymes that hydrolyze the sn-2 acylchain of glycerophospholipids, producing lysophospholipids and free fatty acids. Each PLA 2 enzyme type contributes to specific biological functions based on its expression, subcellular localization, and substrate specificity. Among the PLA 2 superfamily, the cytosolic cPLA 2 enzymes, calcium-independent iPLA 2 enzymes, and secreted sPLA 2 enzymes are implicated in many diseases, but a central issue is the preference for double-bond positions in polyunsaturated fatty acids (PUFAs) occupying the sn-2 position of membrane phospholipids. We demonstrate that each PLA 2 has a unique preference between the specific omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) and the omega-6 arachidonic acid (AA), which are the precursors of most proinflammatory and anti-inflammatory or resolving eicosanoids and related oxylipins. Surprisingly, we discovered that human cPLA 2 selectively prefers AA, whereas iPLA 2 prefers EPA, and sPLA 2 prefers DHA as substrate. We determined the optimal binding of each phospholipid substrate in the active site of each PLA 2 to explain these specificities. To investigate this, we utilized recently developed lipidomics-based LC-MS/MS and GC/MS assays to determine the sn-2 acyl chain specificity in mixtures of phospholipids. We performed s timescale molecular dynamics (MD) simulations to reveal unique active site properties, especially how the precise hydrophobic cavity accommodation of the sn-2 acyl chain contributes to the stability of substrate binding and the specificity of each PLA 2 for AA, EPA, or DHA. This study provides the first comprehensive picture of the unique substrate selectivity of each PLA 2 for omega-3 and omega-6 fatty acids.