Abstract
Adipose triglyceride lipase (ATGL) is the rate-limiting enzyme of lipolysis. ATGL specifically hydrolyzes triacylglycerols (TGs), thereby generating diacylglycerols and free fatty acids. ATGL's enzymatic activity is co-activated by the protein comparative gene identification-58 (CGI-58) and inhibited by the protein G0/G1 switch gene 2 (G0S2). The enzyme is predicted to act through a catalytic dyad (Ser47, Asp166) located within the conserved patatin domain (Ile10-Leu178). Yet, neither an experimentally determined 3D structure nor a model of ATGL is currently available, which would help to understand how CGI-58 and G0S2 modulate ATGL's activity. In this study we determined the minimal active domain of ATGL. This minimal fragment of ATGL could still be activated and inhibited by CGI-58 and G0S2, respectively. Furthermore, we show that this minimal domain is sufficient for protein-protein interaction of ATGL with its regulatory proteins. Based on these data, we generated a 3D homology model for the minimal domain. It strengthens our experimental finding that amino acids between Leu178 and Leu254 are essential for the formation of a stable protein domain related to the patatin fold. Our data provide insights into the structure-function relationship of ATGL and indicate higher structural similarities in the N-terminal halves of mammalian patatin-like phospholipase domain containing proteins, (PNPLA1, -2,- 3 and -5) than originally anticipated.
Highlights
In most organisms, excess energy is stored in form of triacylglycerol (TG) in lipid droplets (LDs)
Activation and inhibition of Adipose triglyceride lipase (ATGL) can be studied in a simplified model system of lipolysis In order to gain more insight into the basic mechanism and structural requirements of ATGL function, its stimulation by CGI58 and inhibition by G0S2, we expanded a simplified experimental setup which was used before to mimic the first step of lipolysis [27]
In vitro assays with mouse ATGL expressed in bacterial cultures harbors the advantages of eliminating possible unidentified factors responsible for reported cell-type dependence and the influence of eukaryotic proteins (e.g. LD associated proteins) from the complex equation of lipolysis
Summary
Excess energy is stored in form of triacylglycerol (TG) in lipid droplets (LDs). Lipolysis is carried out as a hydrolytic cascade of consecutive reactions catalyzed by different lipases [1,2,3,4]. Adipose triglyceride lipase (ATGL) was shown to be the rate-limiting enzyme in this process [1]. It catalyzes the hydrolysis of TG into diacylglycerol (DG) and FA during basal and hormone stimulated lipolysis [1,5,6,7,8]. Patients with mutations in the gene coding for ATGL develop neutral lipid storage disease with myopathy (NLSDM) which is characterized by systemic TG accumulation in multiple tissues and cardiomyopathy [4,9,10]
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