QM/MM Simulations Reveal the Determinants of Carbapenemase Activity in Class A β-lactamases

Ewa I Chudyk, Michael Beer, Michael A L Limb, Charlotte Jones, James Spencer, Marc W Van der Kamp*, Adrian J Mulholland*

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

7 Citations (Scopus)
60 Downloads (Pure)


β-lactam antibiotic resistance in Gram-negative bacteria, primarily caused by β-lactamase enzymes that hydrolyze the β-lactam ring, has become a serious clinical problem. Carbapenems were formerly considered ‘last resort’ antibiotics because they escaped breakdown by most βlactamases, due to slow deacylation of the acyl-enzyme intermediate. However, an increasing number of Gram-negative bacteria now produce β-lactamases with carbapenemase activity: these efficiently hydrolyze the carbapenem β-lactam ring, severely limiting treatment of some bacterial infections. Here, we use quantum mechanics/molecular mechanics (QM/MM) simulations of the deacylation reactions of acyl-enzyme complexes of eight β-lactamases of class A (the most widely distributed β-lactamase group) with the carbapenem meropenem to investigate differences between those inhibited by carbapenems (TEM-1, SHV-1, BlaC, CTX-M-16) and those that hydrolyze them (SFC-1, KPC-2, NMC-A, SME-1). QM/MM molecular dynamics simulationsconfirm the two enzyme groups to differ in the preferred acyl-enzyme orientation: carbapenem-inhibited enzymes favor hydrogen bonding of the carbapenem hydroxyethyl group to the deacylating water (DW). QM/MM simulations of deacylation give activation free energies in good agreement with experimental hydrolysis rates, correctly distinguishing carbapenemases. For the carbapenem-inhibited enzymes activation free energies for deacylation are significantly higher than for the carbapenemases, even when the hydroxyethyl group was restrained to prevent interaction with the DW. Analysis of these simulations, and additional simulations of mutant enzymes, shows how factors including the hydroxyethyl orientation, the active site volume and architecture (conformations of Asn170 and Asn132; organization of the oxyanion hole; and the Cys69-Cys238 disulfide bond) collectively determine catalytic efficiency towards carbapenems.
Original languageEnglish
Pages (from-to)1521-1532
Number of pages12
JournalACS Infectious Diseases
Issue number8
Early online date25 Jul 2022
Publication statusPublished - 12 Aug 2022

Bibliographical note

Funding Information:
A.J.M. and E.C. thank the U.K. Engineering and Physical Science Research Council (EPSRC; grant nos. EP/G007705/1 and EP/M022609/1) for support. M.A.L.L. thanks EPSRC and the Society of Chemical Industry (SCI) for support for a PhD studentship. This work is part of a project that has received funding from the European Research Council under the European Horizon 2020 research and innovation programme (PREDACTED Advanced Grant Agreement no. 101021207) to A.J.M. and J.S. M.W.v.d.K. thanks the U.K. Biotechnology and Biological Sciences Research Council (BBSRC) for funding (BB/M026280/1). J.S. thanks the U.K. Medical Research Council (MRC; U.K.-Canada Team Grant G1100135) for the support. A.J.M. and J.S. thank the MRC (MR/T016035/1). M.B. is supported by the BBSRC-funded South West Biosciences Doctoral Training Partnership [training grant reference BB/T008741/1]. This work was conducted using the computational facilities of the Advanced Computing Research Centre, University of Bristol.

Publisher Copyright:
© 2022 American Chemical Society.


  • Antibiotic resistance
  • carbapenem
  • computational enzymology
  • umbrella sampling
  • electrostatic stabilization
  • ACRC
  • AMR


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