2. Academic Validation
  3. A Two Amino Acid Duplication, L167E168, in the Ω-Loop Drastically Decreases Carbapenemase Activity of KPC-53, a Natural Class A β-Lactamase

A Two Amino Acid Duplication, L167E168, in the Ω-Loop Drastically Decreases Carbapenemase Activity of KPC-53, a Natural Class A β-Lactamase

  • Antimicrob Agents Chemother. 2022 Jun 21;66(6):e0240221. doi: 10.1128/aac.02402-21.
Alessandra Piccirilli 1,  Sabrina Cherubini 1,  Giuseppe Celenza 1,  Gian Maria Rossolini 2 3,  Fabrizia Brisdelli 1,  Bernardetta Segatore 1,  Luigi Principe 4,  Francesco Luzzaro 4,  Lilia Andriani 5,  Gianfranco Amicosante 1,  Mariagrazia Perilli 1
Affiliations

Affiliations

  • 1 Department of Biotechnological and Applied Clinical Sciences, University of L'Aquila, L'Aquila, Italy.
  • 2 Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy.
  • 3 Clinical Microbiology and Virology Unit, Florence Careggi University Hospital, Florence Italy.
  • 4 Clinical Microbiology and Virology Unit, A. Manzoni Hospital, Lecco, Italy.
  • 5 Clinical Pathology and Microbiology Unit, Moriggia Pelascini General Hospital, Gravedona, Italy.
PMID: 35647648 DOI: 10.1128/aac.02402-21
Abstract

KPC-53 Enzyme is a natural KPC variant which showed a duplication of L167E168 residues in the Ω-loop structure. The blaKPC-53 gene was cloned both into pBC-SK and pET-24a vectors, and the recombinant plasmids were transferred by transformation in Escherichia coli competent cells to evaluate the antimicrobial susceptibility and to produce the Enzyme. Compared to KPC-3, the KPC-53 was less stable and showed a dramatic reduction of kcat and kcat/Km versus several β-lactams, in particular carbapenems. Indeed, a 2,000-fold reduction was observed in the kcat values of KPC-53 for imipenem and meropenem. Concerning inhibitors, KPC-53 was susceptible to tazobactam and clavulanic acid but maintained resistance to avibactam. The molecular modeling indicates that the L167E168 duplication in KPC-53 modifies the interactions between residues involved in the catalytic pocket, changing the flexibility of the Ω-loop, which is directly coupled with the catalytic properties of the KPC Enzymes.

Keywords

KPC-53; carbapenemase; serine-β-lactamase.

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