Molecular cloning, expression and characterization of human palmitoyl-protein thioesterase-2

The association of many signal transducing proteins with the plasma membrane often depends upon the dynamic covalent attachment of fatty acids like palmitate. We recently purified and cloned a iysosomal depalmitoylating enzyme, palmitoyl-protein thioesterase (PPT), from bovine brain using H-Ras as substrate. This enzyme was subsequently shown to be defective in the lysosomal storage disease, infantile neuronal ctroid lipofuscino&is. We hereby describe the cloning and expression of another depalmitoyiating enzyme, PPT2. which shares 18% overall identity with PPT1. at the protein level. Northern hybridization revealed a ubiquitous, but low level expression of a predominant 2.0 kb mRNA. The deduced amino acid sequence of PPT2 predicts a protein with 302 amino acids that includes a hydrophobic leader peptide and five potential glycosylation sites. When expressed in simian COS cells, human PPT2 cDNA directs the synthesis of a functional enzyme which is modified by asparagine-linked oligosaccharides and cosediments with lysosomal enzyme markers by Percoll gradient density fractionation. Preliminary experiments indicate, however, that the substrate specificity and tissue dist rib u t ion of PPT2 is different from that of the parent enzyme. PPT1.