Triglyceride-lipase (TGL) is a major fat body lipase in Manduca sexta. The knowledge of how TGL activity is regulated is very limited. A WWE domain, presumably involved in protein-protein interactions, has been previously identified in the N-terminal region of TGL. In this study, we searched for proteins partners that interact with the N-terminal region of TGL. Thirteen proteins were identified by mass spectrometry, and the interaction with four of these proteins was confirmed by immunoblot. The oxidoreductase lipoamide-dehydrogenase (LipDH) and the apolipoprotein components of the lipid transporter, HDLp, were among these proteins. LipDH is the common component of the mitochondrial α-keto acid dehydrogenase complexes whereas HDLp occurs in the hemolymph. However, subcellular fractionation demonstrated that these two proteins are relatively abundant in the soluble fraction of fat body adipocytes. The cofactor lipoate found in typical LipDH substrates was not detected in TGL. However, TGL proved to have critical thiol groups. Additional studies with inhibitors are consistent with the notion that LipDH acting as a diaphorase could preserve the activity of TGL by controlling the redox state of thiol groups. On the other hand, when TG hydrolase activity of TGL was assayed in the presence of HDLp, the production of diacylglycerol (DG) increased. TGL-HDLp interaction could drive the intracellular transport of DG. TGL may be directly involved in the lipoprotein assembly and loading with DG, a process that occurs in the fat body and is essential for insects to mobilize fatty acids. Overall the study suggests that TGL occurs as a multi-protein complex supported by interactions through the WWE domain.
Keywords: Diacylglycerol; Fat body; Lipoamide dehydrogenase; Manduca sexta; Redox; Triglyceride lipase; WWE-domain.
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