title: Discovery of a Specific Inhibitor of Pyomelanin Synthesis in Legionella pneumophila.
abstract: Phenylalanine hydroxylase catalyzes the first step in the synthesis of pyomelanin, a pigment that aids in the acquisition of essential iron in certain bacteria. In this work, we present the development and application of a drug discovery protocol by targeting this enzyme in Legionella pneumophila, the major causative agent of Legionnaires' disease. We employ a combination of high-throughput screening to identify small-molecule binders, enzymatic activity measurements to identify inhibitors in vitro, and the verification of the inhibitory effect in vivo. The most potent inhibitor shows an IC50 value in the low micromolar range and successfully abolishes the synthesis of pyomelanin in L. pneumophila cultures at 10 μM. Thus, this compound represents a novel and effective tool for investigating the role of pyomelanin in the biology and pathogenicity of this organism. Altogether, the results demonstrate a successful pathway for drug development focusing on binding specificity in the initial high-throughput screening steps.
authors.affiliation: Department of Biomedicine, University of Bergen , Bergen, Norway.
authors.forename: Oscar
authors.initials: O
authors.lastname: Aubi
authors.affiliation: Department of Biomedicine, University of Bergen , Bergen, Norway.
authors.forename: Marte I
authors.initials: MI
authors.lastname: Flydal
authors.affiliation: Department of Microbiology and Immunology, Northwestern University Medical School , Chicago, Illinois 60611, United States.
authors.forename: Huaixin
authors.initials: H
authors.lastname: Zheng
authors.affiliation: Department of Biomedicine, University of Bergen , Bergen, Norway.
authors.forename: Lars
authors.initials: L
authors.lastname: Skjærven
authors.affiliation: Department of Chemistry and Centre for Pharmacy, University of Bergen , Bergen, Norway.
authors.forename: Illimar
authors.initials: I
authors.lastname: Rekand
authors.affiliation: The Norwegian Structural Biology Centre (NorStruct), Department of Chemistry, UiT The Arctic University of Norway , Tromsø, Norway.
authors.forename: Hanna-Kirsti S
authors.initials: HK
authors.lastname: Leiros
authors.affiliation: Department of Chemistry and Centre for Pharmacy, University of Bergen , Bergen, Norway.
authors.forename: Bengt Erik
authors.initials: BE
authors.lastname: Haug
authors.affiliation: Department of Microbiology and Immunology, Northwestern University Medical School , Chicago, Illinois 60611, United States.
authors.forename: Nicholas P
authors.initials: NP
authors.lastname: Cianciotto
authors.affiliation: Department of Biomedicine, University of Bergen , Bergen, Norway.
authors.forename: Aurora
authors.initials: A
authors.lastname: Martinez
authors.affiliation: Department of Biomedicine, University of Bergen , Bergen, Norway.
authors.forename: Jarl
authors.initials: J
authors.lastname: Underhaug
mesh_terms: D055808:Drug Discovery; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000494:pharmacology*; D006801:Humans; D007501:Iron / Q000378:metabolism; D016952:Legionella pneumophila / Q000187:drug effects* / Q000201:enzymology* / Q000378:metabolism; D007877:Legionnaires' Disease / Q000188:drug therapy / Q000382:microbiology*; D008024:Ligands; D008543:Melanins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010651:Phenylalanine Hydroxylase / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D008024:Ligands; D008543:Melanins; C023793:pyomelanin; D007501:Iron; D010651:Phenylalanine Hydroxylase