title: Delineation of exoenzyme S residues that mediate the interaction with 14-3-3 and its biological activity.
abstract: 14-3-3 proteins belong to a family of conserved molecules expressed in all eukaryotic cells, which play an important role in a multitude of signaling pathways. 14-3-3 proteins bind to phosphoserine/phosphothreonine motifs in a sequence-specific manner. More than 200 14-3-3 binding partners have been found that are involved in cell cycle regulation, apoptosis, stress responses, cell metabolism and malignant transformation. A phosphorylation-independent interaction has been reported to occur between 14-3-3 and a C-terminal domain within exoenzyme S (ExoS), a bacterial ADP-ribosyltransferase toxin from Pseudomonas aeruginosa. In this study, we have investigated the effect of amino acid mutations in this C-terminal domain of ExoS on ADP-ribosyltransferase activity and the 14-3-3 interaction. Our results suggest that leucine-428 of ExoS is the most critical residue for ExoS enzymatic activity, as cytotoxicity analysis reveals that substitution of this leucine significantly weakens the ability of ExoS to mediate cell death. Leucine-428 is also required for the ability of ExoS to modify the eukaryotic endogenous target Ras. Finally, single amino acid substitutions of positions 426-428 reduce the interaction potential of 14-3-3 with ExoS in vitro.
authors.affiliation: Department of Medical Biosciences/Pathology, Umeå University, Sweden.
authors.forename: Lubna
authors.initials: L
authors.lastname: Yasmin
authors.forename: Anna L
authors.initials: AL
authors.lastname: Jansson
authors.forename: Tooba
authors.initials: T
authors.lastname: Panahandeh
authors.forename: Ruth H
authors.initials: RH
authors.lastname: Palmer
authors.forename: Matthew S
authors.initials: MS
authors.lastname: Francis
authors.forename: Bengt
authors.initials: B
authors.lastname: Hallberg
mesh_terms: D048948:14-3-3 Proteins / Q000502:physiology*; D036002:ADP Ribose Transferases / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology*; D020816:Amino Acid Motifs; D001427:Bacterial Toxins / Q000378:metabolism / Q000494:pharmacology; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D004789:Enzyme Activation; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D007930:Leucine / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D011550:Pseudomonas aeruginosa / Q000201:enzymology; D015398:Signal Transduction / Q000502:physiology
chemical_list: D048948:14-3-3 Proteins; D001427:Bacterial Toxins; D036002:ADP Ribose Transferases; C444820:exoenzyme S; D007930:Leucine