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In vitro depalmitoylation of neurospecific peptides: Implication for infantile neuronal ceroid lipofuscinosis
Author(s) -
Cho Seongeun,
Dawson Philip E.,
Dawson Glyn
Publication year - 2000
Publication title -
journal of neuroscience research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.72
H-Index - 160
eISSN - 1097-4547
pISSN - 0360-4012
DOI - 10.1002/(sici)1097-4547(20000101)59:1<32::aid-jnr5>3.0.co;2-a
Subject(s) - peptide , palmitoylation , biochemistry , thioesterase , biology , chemistry , enzyme , cysteine , biosynthesis
Palmitoyl protein thioesterase 1 (PPT1) removes palmitate from specific cysteine residues in peptides and proteins. We have previously shown that a palmitoylated myelin glycoprotein. Po octapeptide (IRYCWLRR) can be specifically depalmitoylated by PPT1 in vitro (Cho and Dawson [1998] J. Neurochem. 171;323–329). To characterize further the substrate specificity of PPT1, we prepared various palmitoylated oligopeptides, based on palmitoylated sequences from different proteins. A truncated tetrapeptide from Po (RY[palmitoyl]‐CW) was as good a substrate as the octapeptide Po, with optimal activity at pH 4.0. In contrast, other peptide substrates showed marked differences. Thus, the deacylation of GAP‐43 (MLCCMRR), rhodopsin (VTTLCCGKN), and Gα subunit (MGCLGNSK) peptides was more efficient at neutral pH (7.4) than at acidic pH (4.0), with the greatest efficiency toward the Gα peptide (five‐ to sixfold higher than other substrates). Infantile neuronal ceroid lipofuscinosis (INCL) is caused by PPT1 deficiency, and the absence of enzymatic activity was confirmed with GAP‐43 peptide as well as the Po peptide. LA‐N‐5 human neuroblastoma cells overexpressing PPT1 showed increased depalmitoylation of all the peptide substrates, indicating that these peptides are deacylated by PPT1. An amide derivative of a palmitoylated K‐Ras peptide (AcG‐palmitoyl diamino propionate‐VKIKK) acted as an enzyme pseudosubstrate and inhibited PPT1 enzyme activity in a dose‐dependent manner. The peptide itself (AcGCVKIKK) did not affect PPT activity. In summary, PPT1 is able to hydrolyze a range of cysteinyl peptide sequences found in both neuron‐specific and ubiquitous (e.g., Gα) proteins. The inhibitor of PPT1 activity should facilitate the development of a model for INCL and help explain the neuronal death in this disease. J. Neurosci. Res. 59:32–38, 2000 © 2000 Wiley‐Liss, Inc.