Evaluation of the published kinase inhibitor set to identify multiple inhibitors of bacterial ATP-dependent mur ligases

The Mur ligases form a series of consecutive enzymes that participate in the intracellular steps of bacterial peptidoglycan biosynthesis. They therefore represent interesting targets for antibacterial drug discovery. MurC, D, E and F are all ATP-dependent ligases. Accordingly, with the aim being to find multiple inhibitors of these enzymes, we screened a collection of ATP-competitive kinase inhibitors, on <i>Escherichia coli</i> MurC, D and F, and identified five promising scaffolds that inhibited at least two of these ligases. Compounds <b>1</b>, <b>2</b>, <b>4</b> and <b>5</b> are multiple inhibitors of the whole MurC to MurF cascade that act in the micromolar range (IC₅₀, 32–368 µM). NMR-assisted binding studies and steady-state kinetics studies performed on aza-stilbene derivative <b>1</b> showed, surprisingly, that it acts as a competitive inhibitor of MurD activity towards D-glutamic acid, and additionally, that its binding to the D-glutamic acid binding site is independent of the enzyme closure promoted by ATP.