Mechanism of Lipolysis in Insects: Role of Substrate

Abstract

The purpose of this study was to investigate the role of substrate in lipolysis in insects. In insects, lipolysis is the process whereby the neutral lipid triacylglycerol (TAG) is broken down into FFA (free fatty acid) and DAG (diacylglycerol). Lipid droplet (LD), the substrate of lipolysis, is composed of neutral lipids (>95% is TAG), surrounded by a monolayer of amphipathic phospholipids. LD associated proteins are embedded on to the phospholipid monolayer. In order to study the role of substrate in mechanism of lipolysis in insects, we need to analyze the role of lipid droplet associated proteins. Hence we investigated the phosphorylated states lipid droplet associated proteins under AKH stimulated lipolytic conditions. Upon in vivo phosphorylation, the highly phosphorylated protein with an apparent molecular weight of 42-44 kDa protein was subjected to 2-D gel electrophoresis, MALDI-Tof and sequence analysis. The highly phosphorylated protein under AKH stimulated lipolytic conditions was then purified and characterized. Investigation of phosphorylated states of lipid droplet associated proteins under adipokinetic hormone (AKH) stimulated conditions revealed that 42-4kDa doublet protein is the main target of the phosphorylation cascade triggered by AKH, mediated by protein kinase A (PKA). 2-D gel electrophoresis and MALDI-Tof results revealed that the doublet proteins are the isoforms of the same protein. The subsequent sequence analysis revealed that the sequence of the highly phosphorylated protein has a sequence similarity with the lipid storage droplet -1 (LSD-1) protein of Drosophila melanogaster.