Unmasking Crucial Residues in Adipose Triglyceride Lipase (ATGL) for Co-Activation with Comparative Gene Identification-58 (CGI-58)
Author:
Kulminskaya NataliaORCID, Rodriguez Gamez Carlos FranciscoORCID, Hofer PeterORCID, Cerk Ines Kathrin, Dubey NoopurORCID, Viertlmayr Roland, Sagmeister TheoORCID, Pavkov-Keller TeaORCID, Zechner RudolfORCID, Oberer MonikaORCID
Abstract
AbstractLipolysis is an essential metabolic process that releases unesterified fatty acids from neutral lipid stores to maintain energy homeostasis in living organisms. Adipose triglyceride lipase (ATGL) plays a key role in intracellular lipolysis and can be co-activated upon interaction with the protein comparative gene identification-58 (CGI-58). The underlying molecular mechanism of ATGL stimulation by CGI-58 is incompletely understood. Based on analysis of evolutionary conservation, we used site directed mutagenesis to study a C-terminally truncated variant and full-length mouse ATGL providing insights in the protein co-activation on a per-residue level. We identified the region from residues N209-N215 in mouse ATGL as essential for co-activation by mouse CGI-58. ATGL variants with amino-acids exchanges in this region were still able to hydrolyze triacylglycerol at the basal level and to interact with CGI-58, yet could not be activated by CGI-58. Our studies also demonstrate that full-length mouse ATGL showed higher tolerance to specific single amino acid exchanges in the N209-N215 region upon CGI-58 co-activation compared to C-terminally truncated ATGL variants. The region is either directly involved in protein-protein interaction or essential for conformational changes required in the co-activation process. Three-dimensional models of the ATGL/CGI-58 complex with the artificial intelligence software AlphaFold demonstrated that a large surface area is involved in the protein-protein interaction. Mapping important amino acids for co-activation of both proteins, ATGL and CGI-58, onto the 3D model of the complex locates these essential amino acids at the predicted ATGL/CGI-58 interface thus strongly corroborating the significance of these residues in CGI-58 mediated co-activation of ATGL.
Publisher
Cold Spring Harbor Laboratory
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