Metallo-β-Lactamase Inhibitor Phosphonamidate Monoesters

Abstract

Being the second leading cause of death and the leading cause of disability-adjusted life years worldwide, infectious diseases remaincontrary to earlier predictionsa major consideration for the public health of the 21 st century. Resistance development of microbes to antimicrobial drugs constitutes a large part of this devastating problem. The most widely spread mechanism of bacterial resistance operates through the degradation of existing β-lactam antibiotics. Inhibition of metallo-βlactamases is expected to allow the continued use of existing antibiotics, whose applicability is becoming ever more limited. Herein, we describe the synthesis, the metallo-β-lactamase inhibition activity, the cytotoxicity studies, and the NMR spectroscopic determination of the protein binding site of phosphonamidate monoesters. The expression of single-and doublelabeled NDM-1 and its backbone NMR assignment are also disclosed, providing helpful information for future development of NDM-1 inhibitors. We show phosphonamidates to have the potential to become a new generation of antibiotic therapeutics to combat metallo-β-lactamase-resistant bacteria.

Distinct Inhibition Modes of New Delhi Metallo-β-lactamase-1 Revealed by NMR Spectroscopy

“…The 1 H- 15 N TROSY-HSQC and 19 F titration spectra is a relatively sensitive method and provides a wealth of information about protein–inhibitor interaction, such as binding sites, binding constants, and binding modes, and can detect the ligand binding over a wide affinity range, including very weak interactions ( K D > 10 μM). Several NMR studies have been performed to investigate the interactions of NDM-1 with inhibitors. ,, Their results all showed large chemical shift variations from the residues in or near the active sites of di-Zn-NDM-1 upon ligand titrations. 1 H NMR spectroscopy has been reported to real-time monitor the NDM-1 activity in living cells .…”

“…Several NMR studies have been performed to investigate the interactions of NDM-1 with inhibitors. 48,49,60 Their results all showed large chemical shift variations from the residues in or near the active sites of di-Zn-NDM-1 upon ligand titrations. 1 H NMR spectroscopy has been reported to real-time monitor the NDM-1 activity in living cells.…”

“…48 Palica et al have also utilized NMR spectroscopy to determine the binding sites of phosphonamidate monoesters on NDM-1. 49 Here we explore the proteinbased NMR technique to elucidate the inhibition mechanisms of NDM-1 inhibitors and quantitatively characterize the binding affinities of inhibitors. The 1 H-15 N TROSY-HSQC spectrum is sensitive to protein conformation and is regarded as a "fingerprint" of a protein, so it can be used to monitor the global changes of a protein and provide detailed information about the binding and hydrolysis mechanisms.…”

“… 48 Palica et al have also utilized NMR spectroscopy to determine the binding sites of phosphonamidate monoesters on NDM-1. 49 Here we explore the protein-based NMR technique to elucidate the inhibition mechanisms of NDM-1 inhibitors and quantitatively characterize the binding affinities of inhibitors.…”

“…Rivière et al have studied the interactions of several flavonols with NDM-1 using NMR spectroscopy and identified the binding sites on NDM-1 . Palica et al have also utilized NMR spectroscopy to determine the binding sites of phosphonamidate monoesters on NDM-1 . Here we explore the protein-based NMR technique to elucidate the inhibition mechanisms of NDM-1 inhibitors and quantitatively characterize the binding affinities of inhibitors.…”