title: Characterization of a serine hydrolase targeted by acyl-protein thioesterase inhibitors in Toxoplasma gondii.
abstract: In eukaryotic organisms, cysteine palmitoylation is an important reversible modification that impacts protein targeting, folding, stability, and interactions with partners. Evidence suggests that protein palmitoylation contributes to key biological processes in Apicomplexa with the recent palmitome of the malaria parasite Plasmodium falciparum reporting over 400 substrates that are modified with palmitate by a broad range of protein S-acyl transferases. Dynamic palmitoylation cycles require the action of an acyl-protein thioesterase (APT) that cleaves palmitate from substrates and conveys reversibility to this posttranslational modification. In this work, we identified candidates for APT activity in Toxoplasma gondii. Treatment of parasites with low micromolar concentrations of β-lactone- or triazole urea-based inhibitors that target human APT1 showed varied detrimental effects at multiple steps of the parasite lytic cycle. The use of an activity-based probe in combination with these inhibitors revealed the existence of several serine hydrolases that are targeted by APT1 inhibitors. The active serine hydrolase, TgASH1, identified as the homologue closest to human APT1 and APT2, was characterized further. Biochemical analysis of TgASH1 indicated that this enzyme cleaves substrates with a specificity similar to APTs, and homology modeling points toward an APT-like enzyme. TgASH1 is dispensable for parasite survival, which indicates that the severe effects observed with the β-lactone inhibitors are caused by the inhibition of non-TgASH1 targets. Other ASH candidates for APT activity were functionally characterized, and one of them was found to be resistant to gene disruption due to the potential essential nature of the protein.
authors.affiliation: Department of Microbiology and Molecular Medicine, University Medical Center (CMU), University of Geneva, Rue Michel-Servet 1, CH-1211 Geneva, Switzerland.
authors.forename: Louise E
authors.initials: LE
authors.lastname: Kemp
authors.affiliation: Departments of Chemical Biology.
authors.forename: Marion
authors.initials: M
authors.lastname: Rusch
authors.affiliation: Department of Biochemistry, Sciences II, University of Geneva, Quai Ernest Ansermet 30, CH-1211 Geneva, Switzerland.
authors.forename: Alexander
authors.initials: A
authors.lastname: Adibekian
authors.affiliation: Department of Microbiology and Molecular Medicine, University Medical Center (CMU), University of Geneva, Rue Michel-Servet 1, CH-1211 Geneva, Switzerland.
authors.forename: Hayley E
authors.initials: HE
authors.lastname: Bullen
authors.affiliation: Department of Microbiology and Molecular Medicine, University Medical Center (CMU), University of Geneva, Rue Michel-Servet 1, CH-1211 Geneva, Switzerland.
authors.forename: Arnault
authors.initials: A
authors.lastname: Graindorge
authors.affiliation: Department of Parasite Chemotherapy, Swiss Tropical and Public Health Institute, Socinstrasse 57, P. O. Box, CH-4002 Basel, Switzerland; University of Basel, CH-4003 Basel, Switzerland.
authors.forename: Céline
authors.initials: C
authors.lastname: Freymond
authors.affiliation: Department of Parasite Chemotherapy, Swiss Tropical and Public Health Institute, Socinstrasse 57, P. O. Box, CH-4002 Basel, Switzerland; University of Basel, CH-4003 Basel, Switzerland.
authors.forename: Matthias
authors.initials: M
authors.lastname: Rottmann
authors.affiliation: University Montpellier II, CNRS UMR 5235, Montpellier, France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Braun-Breton
authors.affiliation: Departments of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Baumeister
authors.affiliation: Departments of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
authors.forename: Arthur T
authors.initials: AT
authors.lastname: Porfetye
authors.affiliation: Departments of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
authors.forename: Ingrid R
authors.initials: IR
authors.lastname: Vetter
authors.affiliation: Departments of Chemical Biology.
authors.forename: Christian
authors.initials: C
authors.lastname: Hedberg
authors.affiliation: Department of Microbiology and Molecular Medicine, University Medical Center (CMU), University of Geneva, Rue Michel-Servet 1, CH-1211 Geneva, Switzerland. Electronic address: dominique.soldati-favre@unige.ch.
authors.forename: Dominique
authors.initials: D
authors.lastname: Soldati-Favre
mesh_terms: D000595:Amino Acid Sequence; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D006801:Humans; D007783:Lactones / Q000737:chemistry / Q000494:pharmacology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D040681:Structural Homology, Protein; D013869:Thiolester Hydrolases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014122:Toxoplasma / Q000201:enzymology* / Q000235:genetics; D014123:Toxoplasmosis / Q000188:drug therapy / Q000201:enzymology / Q000235:genetics
chemical_list: D004791:Enzyme Inhibitors; D007783:Lactones; D015800:Protozoan Proteins; C402226:LYPLA1 protein, human; C581317:LYPLA2 protein, human; D013869:Thiolester Hydrolases
keywords: Acyl-protein Thioesterase; Enzyme Inhibitors; Hydrolases; Inhibitor; Parasitology; Plasmodium falciparum; Posttranslational Modification; Protein Palmitoylation; Serine Hydrolase; Toxoplasma gondii
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