title: Structures of Pseudomonas aeruginosa β-ketoacyl-(acyl-carrier-protein) synthase II (FabF) and a C164Q mutant provide templates for antibacterial drug discovery and identify a buried potassium ion and a ligand-binding site that is an artefact of the crystal form.
abstract: Bacterial infections remain a serious health concern, in particular causing life-threatening infections of hospitalized and immunocompromised patients. The situation is exacerbated by the rise in antibacterial drug resistance, and new treatments are urgently sought. In this endeavour, accurate structures of molecular targets can support early-stage drug discovery. Here, crystal structures, in three distinct forms, of recombinant Pseudomonas aeruginosa β-ketoacyl-(acyl-carrier-protein) synthase II (FabF) are presented. This enzyme, which is involved in fatty-acid biosynthesis, has been validated by genetic and chemical means as an antibiotic target in Gram-positive bacteria and represents a potential target in Gram-negative bacteria. The structures of apo FabF, of a C164Q mutant in which the binding site is altered to resemble the substrate-bound state and of a complex with 3-(benzoylamino)-2-hydroxybenzoic acid are reported. This compound mimics aspects of a known natural product inhibitor, platensimycin, and surprisingly was observed binding outside the active site, interacting with a symmetry-related molecule. An unusual feature is a completely buried potassium-binding site that was identified in all three structures. Comparisons suggest that this may represent a conserved structural feature of FabF relevant to fold stability. The new structures provide templates for structure-based ligand design and, together with the protocols and reagents, may underpin a target-based drug-discovery project for urgently needed antibacterials.
authors.affiliation: Institut für Pharmazie und Biochemie, Johannes Gutenberg-Universität, Staudinger Weg 5, 55128 Mainz, Germany.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Baum
authors.affiliation: Division of Biological Chemistry and Drug Discovery, College of Life Sciences, University of Dundee, Dundee DD1 4EH, Scotland.
authors.forename: Laura S M
authors.initials: LS
authors.lastname: Lecker
authors.affiliation: Division of Biological Chemistry and Drug Discovery, College of Life Sciences, University of Dundee, Dundee DD1 4EH, Scotland.
authors.forename: Martin
authors.initials: M
authors.lastname: Zoltner
authors.affiliation: Institut für Molekulare Biophysik, Johannes Gutenberg-Universität, Jakob Welder Weg 26, 55128 Mainz, Germany.
authors.forename: Elmar
authors.initials: E
authors.lastname: Jaenicke
authors.affiliation: Department of Medical Biochemistry and Biophysics, Karolinska Institutet, 17 177 Stockholm, Sweden.
authors.forename: Robert
authors.initials: R
authors.lastname: Schnell
authors.affiliation: Division of Biological Chemistry and Drug Discovery, College of Life Sciences, University of Dundee, Dundee DD1 4EH, Scotland.
authors.forename: William N
authors.initials: WN
authors.lastname: Hunter
authors.affiliation: Institut für Pharmazie und Biochemie, Johannes Gutenberg-Universität, Staudinger Weg 5, 55128 Mainz, Germany.
authors.forename: Ruth
authors.initials: R
authors.lastname: Brenk
mesh_terms: D000218:Adamantane / Q000737:chemistry; D020816:Amino Acid Motifs; D062365:Aminobenzoates / Q000737:chemistry; D000813:Anilides / Q000737:chemistry; D000900:Anti-Bacterial Agents / Q000737:chemistry*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics; D001549:Benzamides / Q000737:chemistry; D020134:Catalytic Domain; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D055808:Drug Discovery; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D054889:Fatty Acid Synthase, Type II / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics; D015870:Gene Expression; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011188:Potassium / Q000737:chemistry*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics; D011550:Pseudomonas aeruginosa / Q000737:chemistry* / Q000201:enzymology; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics; D012459:Salicylates / Q000737:chemistry
chemical_list: D062365:Aminobenzoates; D000813:Anilides; D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D001549:Benzamides; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D012459:Salicylates; D054889:Fatty Acid Synthase, Type II; D000218:Adamantane; C510909:platensimycin; D011188:Potassium
keywords: FabF; Gram-negative bacteria; Pseudomonas aeruginosa; fatty-acid biosynthesis; potassium binding; structure-based drug discovery