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The Structural Role of N170 in Substrate-Assisted Deacylation in KPC-2 β-Lactamase

Parwana, Diksha; Gu, Jing; Chen, Shuang; Bethel, Christopher R; Marshall, Emma; Hujer, Andrea M; Bonomo, Robert A; (2024) The Structural Role of N170 in Substrate-Assisted Deacylation in KPC-2 β-Lactamase. Angewandte Chemie International Edition , Article e202317315. 10.1002/anie.202317315. (In press). Green open access

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Abstract

The amino acid substitutions in Klebsiella pneumoniae carbapenemase 2 (KPC-2) that have arisen in the clinic are observed to lead to the development of resistance to ceftazidime-avibactam, a preferred treatment for KPC bearing Gram-negative bacteria. Specific substitutions in the omega loop (R164-D179) results in changes in the structure and function of the enzyme, leading to alterations in substrate specificity, decreased stability, and more recently observed, increased resistance to ceftazidime/avibactam. Using accelerated rare-event sampling well-tempered metadynamics simulations, we explored in detail the structural role of R164 and D179 variants that are described to confer ceftazidime/avibactam resistance. The buried conformation of D179 substitutions produce a pronounced structural disorder in the omega loop - more than R164 mutants, where the crystallographic omega loop structure remains mostly intact. Our findings also reveal that the conformation of N170 plays an underappreciated role impacting drug binding and restricting deacylation. The results further support the hypothesis that KPC-2 D179 variants employ substrate-assisted catalysis for ceftazidime hydrolysis, involving the ring amine of the aminothiazole group to promote deacylation and catalytic turnover. Moreover, the shift in the WT conformation of N170 contributes to reduced deacylation and altered spectrum of enzymatic activity.

Type: Article
Title: The Structural Role of N170 in Substrate-Assisted Deacylation in KPC-2 β-Lactamase
Location: Germany
Open access status: An open access version is available from UCL Discovery
DOI: 10.1002/anie.202317315
Publisher version: http://dx.doi.org/10.1002/anie.202317315
Language: English
Additional information: © 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.
Keywords: β-lactam antibiotics, β-lactamase, metadynamics, substrate-assisted catalysis, enhanced sampling
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > UCL School of Pharmacy
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > UCL School of Pharmacy > Pharma and Bio Chemistry
URI: https://discovery.ucl.ac.uk/id/eprint/10187214
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