Substrate Specificity of OXA-48 after β5-β6 Loop Replacement

Laura Dabos, Agustin Zavala, Rémy A. Bonnin, Oliver Beckstein, Pascal Retailleau, Bogdan I. Iorga, Thierry Naas

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

OXA-48 carbapenemase has rapidly spread in many countries worldwide with several OXA-48-variants being described, differing by a few amino acid (AA) substitutions or deletions, mostly in the β5-β6 loop. While single AA substitutions have only a minor impact on OXA-48 hydrolytic profiles, others with 4 AA deletions result in loss of carbapenem hydrolysis and gain of expanded-spectrum cephalosporin (ESC) hydrolysis. We have replaced the β5-β6 loop of OXA-48 with that of OXA-18, a clavulanic-acid inhibited oxacillinase capable of hydrolyzing ESCs but not carbapenems. The hybrid enzyme OXA-48Loop18 was able to hydrolyze ESCs and carbapenems (although with a lower kcat), even though the β5-β6 loop was longer and its sequence quite different from that of OXA-48. The kinetic parameters of OXA-48Loop18 were in agreement with the MIC values. X-ray crystallography and molecular modeling suggest that the conformation of the grafted loop allows the binding of bulkier substrates, unlike that of the native loop, expanding the hydrolytic profile. This seems to be due not only to differences in AA sequence, but also to the backbone conformation the loop can adopt. Finally, our results provide further experimental evidence for the role of the β5-β6 loop in substrate selectivity of OXA-48-like enzymes and additional details on the structure-function relationship of β-lactamases, demonstrating how localized changes in these proteins can alter or expand their function, highlighting their plasticity.

Original languageEnglish (US)
Pages (from-to)1032-1043
Number of pages12
JournalACS Infectious Diseases
Volume6
Issue number5
DOIs
StatePublished - May 8 2020

Keywords

  • carbapenemase
  • expanded-spectrum cephalosporins
  • oxacillinases
  • β5−β6 loop

ASJC Scopus subject areas

  • Infectious Diseases

Fingerprint Dive into the research topics of 'Substrate Specificity of OXA-48 after β5-β6 Loop Replacement'. Together they form a unique fingerprint.

Cite this