MOLECULAR-CLONING AND EXPRESSION OF PALMITOYL-PROTEIN THIOESTERASE

We have previously reported the purification of a palmitoyl-protein th ioesterase (PPT) from bovine brain that removes palmitate from Ha-Ras (Camp, L. A., and Hofmann, S. L. (1993) J. Biol. Chem. 268, 22566-2257 4). In the current paper, we have isolated bovine and rat cDNA clones encoding PPT. The deduced amino acid sequence of PPT predicts a protei n of 306 amino acids that contains amino acid motifs characteristic of thioesterases: ''Gly-X-Ser-X-Gly'' positioned near the NH, terminus a nd ''Gly-Asp-His'' positioned near the COOH terminus of the protein. T he identity of the PPT cDNA was further confirmed by expression in sim ian COS cells and insect Sf9 cells. Comparison of the DNA and protein sequence data suggests that a hydrophobic NH2-terminal sequence of 27 amino acid residues is removed from the primary translation product. F urthermore, the recombinant protein and the native protein purified fr om bovine brain contain complex asparagine-linked oligosaccharides and a large proportion of the expressed PPT is secreted from COS and Sf9 cells. Thus, while the palmitoyl-protein thioesterase will deacylate i ntracellular palmitoylated proteins such as Ha-Ras and the alpha subun its of heterotrimeric G proteins, the physiologic substrates are likel y to be externally oriented or secreted proteins.