title: Purification and biochemical characterization of a protein-palmitoyl acyltransferase from human erythrocytes.
abstract: Protein palmitoylation involves the post-translational attachment of palmitate in thioester linkage to cysteine residues of proteins. The labile nature of the thioester linkage makes possible the palmitoylation-depalmitoylation cycles that have emerged in recent times as additions to the repertoire of cellular control mechanisms. However, detailed understanding of these cycles has been limited by the lack of knowledge of the transferases and thioesterases likely to be involved. Here, we describe the purification of a protein-palmitoyl acyltransferase (PAT) from human erythrocytes. PAT behaved as a peripheral membrane protein and catalyzed the attachment of palmitate in thioester linkage to the beta-subunit of spectrin. On SDS-polyacrylamide gel electrophoresis, PAT appeared as a 70-kDa polypeptide. Antibody against this polypeptide could immunodeplete PAT activity from the crude extract, confirming the assignment of the 70-kDa polypeptide as PAT. PAT-mediated spectrin palmitoylation could be inhibited by nonradioactive palmitoyl-, myristoyl-, or stearoyl-CoA. The apparent Km for palmitoyl-CoA was 16 microM.
authors.affiliation: Department of Chemistry, Bose Institute, 93/1 Acharya Prafulla Chandra Road, Calcutta 700 009, India.
authors.forename: A K
authors.initials: AK
authors.lastname: Das
authors.forename: B
authors.initials: B
authors.lastname: Dasgupta
authors.forename: R
authors.initials: R
authors.lastname: Bhattacharya
authors.forename: J
authors.initials: J
authors.lastname: Basu
mesh_terms: D000215:Acylation; D000217:Acyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D010168:Palmitates / Q000378:metabolism*; D010171:Palmitoyl Coenzyme A / Q000378:metabolism; D010449:Peptide Mapping; D011499:Protein Processing, Post-Translational; D013049:Spectrin / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D010168:Palmitates; D013049:Spectrin; D010171:Palmitoyl Coenzyme A; D000217:Acyltransferases; C105599:HHAT protein, human