TY - JOUR
AU - Black, Michael T.
AB - Mechanistic Studies of the Inactivation of TEM-1 and P99 by NXL104, a Novel Non-β-Lactam β-Lactamase Inhibitor ▿ Thérèse Stachyra 1 , Marie-Claude Péchereau 1 , Jean-Michel Bruneau 1 , Monique Claudon 1 , Jean-Marie Frère 2 , Christine Miossec 1 , Kenneth Coleman 1 and Michael T. Black 1 , * 1 Novexel SA, Parc Biocitech, 102 Avenue Gaston Roussel, 93230 Romainville, France 2 CIP/Enzymology, Université de Liège, Institute of Chemistry B6, Sart-Tilman B4000 Liège, Belgium ABSTRACT NXL104 is a potent inhibitor of class A and C serine β-lactamases, including KPC carbapenemases. Native and NXL104-inhibited TEM-1 and P99 β-lactamases analyzed by liquid chromatography-electrospray ionization-time of flight mass spectrometry revealed that the inactivated enzymes formed a covalent adduct with NXL104. The principal inhibitory characteristics of NXL104 against TEM-1 and P99 β-lactamases were determined, including partition ratios, dissociation constants ( K ), rate constants for deactivation ( k 2 ), and reactivation rates. NXL104 is a potent inhibitor of TEM-1 and P99, characterized by high carbamylation efficiencies ( k 2 / K of 3.7 × 10 5 M −1 s −1 for TEM-1 and 1 × 10 4 M −1 s −1 for P99) and slow decarbamylation. Complete loss of β-lactamase activity was obtained at a 1/1 enzyme/NXL104 ratio, with a k 3 value (rate constant for formation of product and free enzyme) close to zero for TEM-1 and P99. Fifty percent inhibitory concentrations (IC 50 s) were evaluated on selected β-lactamases, and NXL104 was shown to be a very potent inhibitor of class A and C β-lactamases. IC 50 s obtained with NXL104 (from 3 nM to 170 nM) were globally comparable on the β-lactamases CTX-M-15 and SHV-4 with those obtained with the comparators (clavulanate, tazobactam, and sulbactam) but were far lower on TEM-1, KPC-2, P99, and AmpC than those of the comparators. In-depth studies on TEM-1 and P99 demonstrated that NXL104 had a comparable or better affinity and inactivation rate than clavulanate and tazobactam and in all cases an improved stability of the covalent enzyme/inhibitor complex.
TI - Mechanistic Studies of the Inactivation of TEM-1 and P99 by NXL104, a Novel Non-β-Lactam β-Lactamase Inhibitor
JF - Antimicrobial Agents and Chemotherapy
DO - 10.1128/AAC.00568-10
DA - 2010-12-01
UR - https://www.deepdyve.com/lp/american-society-for-microbiology/mechanistic-studies-of-the-inactivation-of-tem-1-and-p99-by-nxl104-a-la0vxWtdX0
SP - 5132
VL - 54
IS - 12
DP - DeepDyve
ER -