Discovery of novel benzene 1,3-dicarboxylic acid inhibitors of bacterial MurD and MurE ligases by structure-based virtual screening approach Andrej Perdih a , Andreja Kovac ˇ b , Gerhard Wolber c,d , Didier Blanot e , Stanislav Gobec b , Tom Solmajer a, * a National Institute of Chemistry, Hajdrihova 19, 1001 Ljubljana, Slovenia b Faculty of Pharmacy, University of Ljubljana, Aškerc ˇeva 7, 1000 Ljubljana, Slovenia c Inte:Ligand GmbH, Mariahilferstrasse 74B/11, 1070 Vienna, Austria d Institute of Pharmacy and Center for Molecular Biosciences Innsbruck (CMBI), University of Innsbruck, Innrain 52c, 6020 Innsbruck, Austria e Enveloppes Bactériennes et Antibiotiques, IBBMC, UMR 8619 CNRS, Univ Paris-Sud, 91405 Orsay, France article info Article history: Received 16 February 2009 Revised 25 March 2009 Accepted 27 March 2009 Available online 1 April 2009 Keywords: MurD and MurE enzymes Virtual screening Three-dimensional structure-based pharmacophores Molecular docking Antibacterial agents Drug design abstract The peptidoglycan biosynthetic pathway provides an array of potential targets for antibacterial drug design, attractive especially with respect to selective toxicity. Within this pathway, the members of the Mur ligase family are considered as promising emerging targets for novel antibacterial drug design. Based on the available MurD crystal structures co-crystallised with N-sulfonyl glutamic acid inhibitors, a virtual screening campaign was performed, combining three-dimensional structure-based pharmaco- phores and molecular docking calculations. A novel class of glutamic acid surrogates—benzene 1,3-dicar- boxylic acid derivatives—were identified and compounds 14 and 16 found to possess dual MurD and MurE inhibitory activity. Ó 2009 Elsevier Ltd. All rights reserved. The increasing incidence of bacterial resistance to most of the available antibiotics has rendered the discovery of novel efficacious antibacterial agents imperative, suggesting the consideration of novel, previously unexploited targets. 1,2 An essential component unique to prokaryotic cells—the peptidoglycan—is traditionally a target of choice with respect to selective toxicity. 3–5 The biosynthe- sis of the peptidoglycan is a complex multistage process divided into an early stage of intracellular assembly of the UDP-MurNAc- pentapeptide, followed by a translocation step on the outercellular side and final incorporation into the nascent biopolymer. 3–5 Prop- erly constructed peptidoglycan provides rigidity, flexibility and strength that are necessary for bacterial cells to grow and divide, while withstanding high internal osmotic pressure. 6 The four members of the ADP-forming bacterial ligase family (the synthesis of the peptide bond is coupled with a concurrent hydrolysis of the ATP molecules to ADP and P i )—MurC, MurD, MurE and MurF—are involved in the intracellular phase of peptidoglycan assembly, catalysing the synthesis of the peptide moiety by con- secutive addition of L-Ala, D-Glu, meso-A 2 pm (or L-Lys) and D-Ala- D-Ala to the UDP-N-acetylmuramic acid (UDP-MurNAc), the start- ing UDP-precursor. 3,7 The Mur ligase family is considered a prime example of modular structure in protein architecture, with molecules made up of three domains or modules allowing for molecular recognition of a spe- cific UDP-substrate. 3,7 The three-domain structure of the family comprises an N-terminal domain responsible for binding the UDP-substrate, a central domain bearing resemblance to the ATP- binding domains of a number of ATP- or GTP-ases and a C-terminal domain, which is involved in the binding of the incoming amino acid. 7 Structural investigations of the Mur ligase family have re- sulted in the identification of different protein conformations, making the whole family a complex collection of dynamic protein targets. 7,8 All Mur ligases presumably act through an analogous sequential enzymatic mechanism, as corroborated by structural, 9 biochemical 10 and computational studies. 11 In the proposed mech- anism, the bound UDP-precursor 1 initially reacts with the ATP molecule yielding an acyl-phosphate intermediate, 10 which, fol- lowing addition of the incoming amino acid, affords the tetrahedral reaction intermediate 2. 9,11 Finally, the dissociation of the phos- phate group results in a new UDP-precursor elongated by the con- densed amino acid 3. 9 The attraction of the Mur ligase family as a collection of emerg- ing drug targets has resulted in the design of several ligase inhib- itors. 3,12–17 A frequently used starting point in the design of MurD (UDP-N-acetylmuramoyl-L-alanyl:D-glutamate ligase) 0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmcl.2009.03.141 * Corresponding author. Tel.: +386 0 1 4760 277; fax: +386 0 1 4760 300. E-mail address: tom.solmajer@ki.si (T. Solmajer). Bioorganic & Medicinal Chemistry Letters 19 (2009) 2668–2673 Contents lists available at ScienceDirect Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl