Acyl-protein thioesterases are enzymes that cleave off lipid modifications on proteins, located on the sulfur atom of cysteine residues linked via a thioester bond.^[1] Acyl-protein thioesterases are part of the α/β hydrolase superfamily of proteins and have a conserved catalytic triad.^[2] For that reason, acyl-protein thioesterases are also able to hydrolyze oxygen-linked ester bonds.

Acyl-protein thioesterases are involved in the depalmitoylation of proteins, meaning they cleave off palmitoyl modifications on proteins' cysteine residues. Cellular targets include trimeric G-alpha proteins,^[3] ion channels^[4] and GAP-43.^[5] Moreover, human acyl-protein thioesterases 1 and 2 have been identified as major components in controlling the palmitoylation cycle of the oncogene Ras.^[6][7] Depalmitoylation of Ras by acyl-protein thioesterases potentially reduces Ras' affinity to endomembranes, allowing it to be palmitoylated again at the Golgi apparatus and to be directed to the plasma membrane. Acyl-protein thioesterases, therefore, are thought to correct potential mislocalization of Ras.

Currently fully validated human acyl-protein thioesterases are APT1^[8] and APT2^[9] which share 66% sequence homology.^[10] Additionally there are a handful of putative acyl-protein thioesterases reported, including the ABHD17 enzyme family.^[11][12] In the lysosome, PPT1 of the palmitoyl protein thioesterase family has similar enzymatic activity as acyl-protein thioesterases.

Acyl-protein thioesterases feature 3 major structural components that determine protein function and substrate processing: 1. A conserved, classical catalytic triad to break ester and thioester bonds;^[2] 2. A long hydrophobic substrate tunnel to accommodate the palmitoyl moiety, as identified in the crystal structures of human acyl-protein thioesterase 1,^[2] human acyl-protein thioesterase 2^[13] and Zea mays acyl-protein thioesterase 2;^[14] 3. A lid-loop that covers the catalytic site, is highly flexible and is a main factor determining the enzyme's product release rate.^[14]

The involvement in controlling the localization of the oncogene Ras has made acyl-protein thioesterases potential cancer drug targets.^[15] Inhibition of acyl-protein thioesterases is believed to increase mislocalization of Ras at the cell's membranes, eventually leading to a collapse of the Ras cycle. Inhibitors for acyl-protein thioesterases have been specifically targeting the hydrophobic substrate tunnel,^[16][13] the catalytic site serine^[17] or both.^[18]

Current approaches to study the biological activity of Acyl-protein Thioesterases include proteomics, monitoring the trafficking of microinjected fluorescent substrates,^[19][7] the use of cell-permeable substrate mimetics,^[20] and cell permeable small molecule fluorescent chemical tools.^{[21][22][23][24]}