<i>Trypanosoma brucei</i> Acyl-Protein Thioesterase-like (TbAPT-L) Is a Lipase with Esterase Activity for Short and Medium-Chain Fatty Acids but Has No Depalmitoylation Activity
Robert W. B. Brown,
Aabha I. Sharma,
Miguel Rey Villanueva,
Xiaomo Li,
Ouma Onguka,
Leeor Zilbermintz,
Helen Nguyen,
Ben A. Falk,
Cheryl L. Olson,
Joann M. Taylor,
Conrad L. Epting,
Rahul S. Kathayat,
Neri Amara,
Bryan C. Dickinson,
Matthew Bogyo,
David M. Engman
Affiliations
Robert W. B. Brown
Departments of Pathology, Microbiology-Immunology and Pediatrics, Northwestern University, Chicago, IL 60611, USA
Aabha I. Sharma
Departments of Pathology, Microbiology-Immunology and Pediatrics, Northwestern University, Chicago, IL 60611, USA
Miguel Rey Villanueva
Department of Pathology and Laboratory Medicine, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
Xiaomo Li
Departments of Pathology, Microbiology-Immunology and Pediatrics, Northwestern University, Chicago, IL 60611, USA
Ouma Onguka
Departments of Pathology and Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA
Leeor Zilbermintz
Department of Pathology and Laboratory Medicine, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
Helen Nguyen
Department of Pathology and Laboratory Medicine, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
Ben A. Falk
Department of Pathology and Laboratory Medicine, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
Cheryl L. Olson
Departments of Pathology, Microbiology-Immunology and Pediatrics, Northwestern University, Chicago, IL 60611, USA
Joann M. Taylor
Departments of Pathology, Microbiology-Immunology and Pediatrics, Northwestern University, Chicago, IL 60611, USA
Conrad L. Epting
Departments of Pathology, Microbiology-Immunology and Pediatrics, Northwestern University, Chicago, IL 60611, USA
Rahul S. Kathayat
Department of Chemistry, The University of Chicago, Chicago, IL 60637, USA
Neri Amara
Departments of Pathology and Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA
Bryan C. Dickinson
Department of Chemistry, The University of Chicago, Chicago, IL 60637, USA
Matthew Bogyo
Departments of Pathology and Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA
David M. Engman
Departments of Pathology, Microbiology-Immunology and Pediatrics, Northwestern University, Chicago, IL 60611, USA
Dynamic post-translational modifications allow the rapid, specific, and tunable regulation of protein functions in eukaryotic cells. S-acylation is the only reversible lipid modification of proteins, in which a fatty acid, usually palmitate, is covalently attached to a cysteine residue of a protein by a zDHHC palmitoyl acyltransferase enzyme. Depalmitoylation is required for acylation homeostasis and is catalyzed by an enzyme from the alpha/beta hydrolase family of proteins usually acyl-protein thioesterase (APT1). The enzyme responsible for depalmitoylation in Trypanosoma brucei parasites is currently unknown. We demonstrate depalmitoylation activity in live bloodstream and procyclic form trypanosomes sensitive to dose-dependent inhibition with the depalmitoylation inhibitor, palmostatin B. We identified a homologue of human APT1 in Trypanosoma brucei which we named TbAPT-like (TbAPT-L). Epitope-tagging of TbAPT-L at N- and C- termini indicated a cytoplasmic localization. Knockdown or over-expression of TbAPT-L in bloodstream forms led to robust changes in TbAPT-L mRNA and protein expression but had no effect on parasite growth in vitro, or cellular depalmitoylation activity. Esterase activity in cell lysates was also unchanged when TbAPT-L was modulated. Unexpectedly, recombinant TbAPT-L possesses esterase activity with specificity for short- and medium-chain fatty acid substrates, leading to the conclusion, TbAPT-L is a lipase, not a depalmitoylase.
