Contributors: Armand-Frappier Santé Biotechnologie Research Centre (INRS-AFSB); Institut National de la Recherche Scientifique Québec (INRS)-Pasteur Network (Réseau International des Instituts Pasteur); Groupe de Recherche Axé sur la Structure des Protéines (GRASP); McGill University = Université McGill Montréal, Canada; PROTEO, The Quebec Network for Research on Protein Function, Engineering, and Applications; Institut National de la Recherche Scientifique Québec (INRS)-Pasteur Network (Réseau International des Instituts Pasteur)-Institut National de la Recherche Scientifique Québec (INRS)-Pasteur Network (Réseau International des Instituts Pasteur)-Université de Sherbrooke (UdeS)-Université Laval Québec (ULaval)-McGill University = Université McGill Montréal, Canada -University of Ottawa Ottawa -Université du Québec à Trois-Rivières (UQTR)-Université de Montréal (UdeM)-TransBiotech, Lévis-Concordia University Montreal -Université du Québec à Montréal = University of Québec in Montréal (UQAM); This work was supported by Natural Sciences and Engineering Research Council Discovery grant RGPIN 402623-2011 (to N.D.), CIHR operating grant MOP-97888 (to E.D.), and a FRQS Research Scholar Junior 1 Career Award (to N.D.). N.D. also acknowledges support from the FRQNT Strategic Cluster “Regroupement Québécois de Recherche sur la Fonction, la Structure et l'Ingénierie des Protéines” (PROTEO) and the FRQS Strategic Cluster “Groupe de Recherche Axé sur la Structure des Protéines” (GRASP). E.D. holds a Canada Research Chair in sociomicrobiology. B.F. is the recipient of a “Fondation Universitaire Armand-Frappier de l'INRS” postdoctoral fellowship.
نبذة مختصرة : International audience ; Pseudomonas aeruginosa is an important opportunistic human pathogen that can establish bacterial communication by synchronizing the behavior of individual cells in a molecular phenomenon known as "quorum sensing". Through an elusive mechanism involving gene products of the pqs operon, the PqsE enzyme is absolutely required for the synthesis of extracellular phenazines, including the toxic blue pigment pyocyanin, effectively allowing cells to achieve full-fledged virulence. Despite several functional and structural attempts at deciphering the role of this relevant enzymatic drug target, no molecular function has yet been ascribed to PqsE. In the present study, we report a series of alanine scanning experiments aimed at altering the biological function of PqsE, allowing us to uncover key amino acid positions involved in the molecular function of this enzyme. We use sequence analysis and structural overlays with members of homologous folds to pinpoint critical positions located in the vicinity of the ligand binding cleft and surrounding environment, revealing the importance of a unique C-terminal α-helical motif in the molecular function of PqsE. Our results suggest that the active site of the enzyme involves residues that extend further into the hydrophobic core of the protein, advocating for a lid-like movement of the two terminal helices. This information should help design virtual libraries of PqsE inhibitors, providing means to counter P. aeruginosa virulence acquisition and helping to reduce nosocomial infections.
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