Mechanistic Insights into β-Lactamase-Catalysed Carbapenem Degradation Through Product Characterisation

Christopher T. Lohans, Emily I. Freeman, Emma van Groesen, Catherine L. Tooke, Philip Hinchliffe, James Spencer, Jürgen Brem, Christopher J. Schofield

Research output: Contribution to journalArticle (Academic Journal)

3 Citations (Scopus)
114 Downloads (Pure)

Abstract

β-Lactamases are a major threat to the clinical use of carbapenems, which are often antibiotics of last resort. Despite this, the reaction outcomes and mechanisms by which β-lactamases degrade carbapenems are still not fully understood. The carbapenem bicyclic core consists of a β-lactam ring fused to a pyrroline ring. Following β-lactamase-mediated opening of the β-lactam, the pyrroline may interconvert between an enamine (2-pyrroline) form and two epimeric imine (1-pyrroline) forms; previous crystallographic and spectroscopic studies have reported all three of these forms in the contexts of hydrolysis by different β-lactamases. As we show by NMR spectroscopy, the serine β-lactamases (KPC-2, SFC-1, CMY-10, OXA-23, and OXA-48) and metallo-β-lactamases (NDM-1, VIM-1, BcII, CphA, and L1) tested all degrade carbapenems to preferentially give the Δ2 (enamine) and/or (R)-Δ1 (imine) products. Rapid non-enzymatic tautomerisation of the Δ2 product to the (R)-Δ1 product prevents assignment of the nascent enzymatic product by NMR. The observed stereoselectivity implies that carbapenemases control the form of their pyrroline ring intermediate(s)/product(s), thereby preventing pyrroline tautomerisation from inhibiting catalysis.

Original languageEnglish
Article number13608
Number of pages9
JournalScientific Reports
Volume9
DOIs
Publication statusPublished - 20 Sep 2019

Keywords

  • Antibiotics
  • Antimicrobial resistance
  • Enzyme mechanisms
  • hydrolases

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