A Novel inhibitor of Lipoxygenase: a Patatin‐like Phospholipase Domain‐containing 2 (PNPLA2) Protein

Abstract

Lipoxygenases (LOX) are enzymes responsible for the metabolism of arachidonic acid and other polyunsaturated fatty acids, thereby contributing to the generation of reactive oxygen species (ROS) under oxidative stress. Pigment epithelium‐derived factor receptor (PEDF‐R), a PNPLA2 protein, contains two peptide regions E5b (I193–L232) and P1 (T210‐L249) that bind PEDF. Preliminary experiments showed that these peptides inhibited the coupled PLA/LOX enzymatic reaction and protected ARPE‐19 cells from oxidative damage. Here we investigated whether the above observation was due to direct binding and inhibition of lipoxygenase activity by the peptides. Peptide affinity chromatography demonstrated that soybean lipoxygenase V specifically bound to synthetic peptides E5b and P1 immobilized to agarose. Among several different buffers, optimal binding was observed with the LOX enzymatic reaction buffer. Spectrophotometric assays showed that E5b or P1 inhibited LOX activity in a concentration dependent manner. Interestingly, this inhibition was blocked when E5b was mixed with Fl‐PEDF, suggesting that Fl‐PEDF captures the peptide making it unavailable for LOX inhibition. Our results support the idea that the PEDF‐R via binding and inhibiting a mamalian lipoxygenase has protective effect from oxidative damage. Supported by NIHNEI‐IRP.