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Benzothioxalone derivatives as novel inhibitors of UDP-N-acetylglucosamine enolpyruvyl transferases (MurA and MurZ)

Miller, Keith and Dunsmore, Colin and Leeds, Jennifer and Patching, Simon and Sachdeva, Meena and Blake, Katy and Stubbings, William and Simmons, Katie and Henderson, Peter and De Los Angeles, Joseph and Fishwick, Colin and Chopra, Ian (2010) Benzothioxalone derivatives as novel inhibitors of UDP-N-acetylglucosamine enolpyruvyl transferases (MurA and MurZ). Journal of Antimicrobial Chemotherapy, 65 (12). pp. 2566-2573. ISSN 1460-2091

Abstract

Objectives: We sought to identify and characterize new inhibitors of MurA and MurZ which are enzymes involved in the early stages of bacterial peptidoglycan synthesis.
Methods: A library of approximately 650,000 compounds was screened for inhibitors of Escherichia coli MurA in an endpoint assay measuring release of inorganic phosphate from phosphoenolpyruvate. Hits were validated by determining the concentrations required for 50% inhibition (IC50) of MurA from E. coli and MurA/MurZ from Staphylococcus aureus. The mode of action of selected inhibitors was explored by examining the reversibility of MurA inhibition, the binding of a radiolabelled inhibitor to MurA proteins and through docking studies. Inhibitors were further characterized by determining their anti-bacterial activities against E. coli and S. aureus
Results: Benzothioxalone derivatives were identified which inhibited MurA from E. coli and MurA/MurZ from S. aureus with IC50 values between 0.25 and 51 M. Several inhibitors also exhibited activity against S. aureus with MICs in the range 4 – 128 mg/L. Inhibition of MurA was irreversible and a radiolabelled inhibitor from this compound class displayed stoichiometric binding to the enzyme, which was displaced by dithiothreitol. Binding was undetectable with a C115D mutant MurA protein.
Conclusions: The results suggests a mode of action for the benzothioxalones that involves the formation of a disulfide bond with MurA/MurZ, via attack from an active site cysteine on the thioxalone ring carbonyl group, followed by ring opening to yield an S-acylated protein. The proposed covalent mode of action may prove useful in the design of new antibacterial agents.

Item Type: Article
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Additional Information: author can archive post-print (ie final draft post-refereeing); Publisher's version/PDF cannot be used
Keywords: peptidoglycan synthesis; Mur enzymes; drug targets
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Date Deposited: 13 Oct 2015 13:16
Last Modified: 13 Oct 2015 13:16
URI: https://oak.novartis.com/id/eprint/2997

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