2. Academic Validation
  3. Restricted Rotational Flexibility of the C5α-Methyl-Substituted Carbapenem NA-1-157 Leads to Potent Inhibition of the GES-5 Carbapenemase

Restricted Rotational Flexibility of the C5α-Methyl-Substituted Carbapenem NA-1-157 Leads to Potent Inhibition of the GES-5 Carbapenemase

  • ACS Infect Dis. 2024 Mar 21. doi: 10.1021/acsinfecdis.3c00683.
Nichole K Stewart 1,  Marta Toth 1,  Pojun Quan 2,  Michael Beer 3 4,  John D Buynak 2,  Clyde A Smith 5 6,  Sergei B Vakulenko 1
Affiliations

Affiliations

  • 1 Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, Indiana 46556, United States.
  • 2 Department of Chemistry, Southern Methodist University, Dallas, Texas 75275, United States.
  • 3 School of Cellular and Molecular Medicine, University of Bristol, Bristol BS8 1TD, U.K.
  • 4 Centre for Computational Chemistry, School of Chemistry, University of Bristol, Bristol BS8 1TS, U.K.
  • 5 Stanford Synchrotron Radiation Lightsource, Stanford University, Menlo Park, California 94025, United States.
  • 6 Department of Chemistry, Stanford University, Stanford, California 94305, United States.
PMID: 38511828 DOI: 10.1021/acsinfecdis.3c00683
Abstract

Carbapenem Antibiotics are used as a last-resort treatment for infections caused by multidrug-resistant bacteria. The wide spread of carbapenemases in Gram-negative bacteria has severely compromised the utility of these drugs and represents a serious public health threat. To combat carbapenemase-mediated resistance, new antimicrobials and inhibitors of these enzymes are urgently needed. Here, we describe the interaction of the atypically C5α-methyl-substituted carbapenem, NA-1-157, with the GES-5 carbapenemase. MICs of this compound against Escherichia coli, Klebsiella pneumoniae, and Acinetobacter baumannii producing the Enzyme were reduced 4-16-fold when compared to MICs of the commercial carbapenems, reaching clinically sensitive breakpoints. When NA-1-157 was combined with meropenem, a strong synergistic effect was observed. Kinetic and ESI-LC/MS studies demonstrated that NA-1-157 is a potent inhibitor of GES-5, with a high inactivation efficiency of (2.9 ± 0.9) × 105 M-1 s-1. Acylation of GES-5 by NA-1-157 was biphasic, with the fast phase completing within seconds, and the slow phase taking several hours and likely proceeding through a reversible tetrahedral intermediate. Deacylation was extremely slow (k3 = (2.4 ± 0.3) × 10-7 s-1), resulting in a residence time of 48 ± 6 days. MD simulation of the GES-5-meropenem and GES-5-NA-1-157 acyl-enzyme complexes revealed that the C5α-methyl group in NA-1-157 sterically restricts rotation of the 6α-hydroxyethyl group preventing ingress of the deacylating water into the vicinity of the scissile bond of the acyl-enzyme intermediate. These data demonstrate that NA-1-157 is a potent irreversible inhibitor of the GES-5 carbapenemase.

Keywords

antibiotic resistance; carbapenem; carbapenemase; crystal structure; inhibitor; kinetics.

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