An optimized protocol for identifying S-acylated proteins in Dictyostelium discoideum

S-palmitoylation is a reversible post-translational modification that adds palmitic acid onto cysteine residues of proteins through the formation of a thioester bond. The reaction is catalyzed by protein acyl transferases (PATs) and reversed by acyl protein thioesterases (APTs), also known as S-depalmitoylases. Here, we optimized acyl resin-assisted capture (acyl-RAC) for identifying S-palmitoylated proteins in the model eukaryote Dictyostelium discoideum. Using this optimized protocol and western blotting, we revealed S-palmitoylated proteins in D. discoideum including calcium-dependent cell adhesion protein A (CadA), calreticulin (CalR), and glucose-regulated protein 78 (Grp78). Overall, this work establishes acyl-RAC as a tool for studying S-palmitoylation in D. discoideum and reveals a subset of S-palmitoylated proteins in this model organism that can be further studied.