Abstract
A series of optimized sulfonamide derivatives was recently reported as novel inhibitors of UDP-N-acetylmuramoyl-L-alanine:D-glutamate ligase (MurD). These are based on naphthalene-N-sulfonyl-D-glutamic acid and have the D-glutamic acid replaced with rigidified mimetics. Here we have defined the binding site of these novel ligands to MurD using 1H/13C heteronuclear single quantum correlation. The MurD protein was selectively 13C-labeled on the methyl groups of Ile (δ1 only), Leu and Val, and was isolated and purified. Crucial Ile, Leu and Val methyl groups in the vicinity of the ligand binding site were identified by comparison of chemical shift perturbation patterns among the ligands with various structural elements and known binding modes. The conformational and dynamic properties of the bound ligands and their binding interactions were examined using the transferred nuclear Overhauser effect and saturation transfer difference. In addition, the binding mode of these novel inhibitors was thoroughly examined using unrestrained molecular dynamics simulations. Our results reveal the complex dynamic behavior of ligand–MurD complexes and its influence on ligand–enzyme contacts. We further present important findings for the rational design of potent Mur ligase inhibitors.
Highlights
The increasing rate of bacterial resistance against available antibacterial agents is becoming a serious threat to our society
The Mur ligases are essential intracellular bacterial enzymes that are involved in the biosynthesis of peptidoglycan precursors and represent attractive targets for the development of novel antibiotics
MurD was titrated with eleven naphthalene-N-sulfonyl derivatives (1, 1a, 2a, 5a, 6a, and 1b–6b)
Summary
The increasing rate of bacterial resistance against available antibacterial agents is becoming a serious threat to our society. The Mur ligases are essential intracellular bacterial enzymes that are involved in the biosynthesis of peptidoglycan precursors and represent attractive targets for the development of novel antibiotics. The Mur ligase family comprises enzymes UDP-Nacetylmuramate:L-alanine ligase (MurC), UDP-N-acetylmuramoyl-L-alanine:D-glutamate ligase (MurD), UDP-N-acetylmuramoyl-L-alanyl-D-glutamate:meso-diaminopimelate ligase (MurE), and UDP-N-acetylmuramoyl-L-alanyl-c-D-glutamyl-meso-diaminopimelate:D-alanyl-D-alanine ligase (MurF) These enzymes consecutively add L-Ala (MurC), D-Glu (MurD), meso-2,6diaminopimelic acid (or L-Lys in most Gram-positive bacteria) (MurE), and D-Ala- D-Ala (MurF) to the nucleotide precursor uridine 59-diphosphate-N-acetylmuramic acid (UDP-MurNAc) [3]. A high-energy, tetrahedral intermediate is produced that yields the nucleotide products, ADP and inorganic phosphate [3]
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