Activation of phospholipase A2 by prostaglandin in vitro

Prostaglandins are a diverse family of biological active molecules that are synthesized after liberation of arachnidonic or linolenic acid from the plasma membrane by phospholipase A2 (PLA2). Specific prostaglandins may be pro-inflammatory or anti-inflammatory due to a poorly understood biochemical equilibrium. Some of the anti-inflammatory prostaglandins namely, prostaglandin A1 (PGA1) and prostaglandin E1 (PGE1) have a cyclopentenone moiety that can react and modify a protein's activity. These two prostaglandins are electrophilic reactive lipid species and are formed as a result of the reaction cascade initiated by PLA2. It was of interest to study the interaction with these prostaglandins as they could either amplify or block this enzyme's activity. We found that the former is true initially as there is a shorter time to activate the protein on the lipid membrane and an overall increase in hydrolysis was observed when PGA1 and PGE1 prostaglandin was added with PLA2 and liposomes. The interfacial activation model was further explored in which there is a modification of the enzyme rather than a modifcation of the substrate. However, after a time the protein was shown to form amyloid like fibrils thereby blocking further hydrolysis. The fibrillization kinetics in the presence of the one of the prostaglandins was monitored using thioflavin T (ThT) and the resulting fibrils were characterized using transmission electron microscopy (TEM) and atomic force microscopy (AFM). Modification of PLA2 by these prostaglandins leading to amyloid like fibrils gives an additional perspective of control of the interfacial activation mechanism of this enzyme.