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Photo Lynn Kamerlin

Lynn Kamerlin

Professor

Photo Lynn Kamerlin

Targeting MarA N-terminal domain dynamics to prevent DNA binding

Author

  • Marina Corbella
  • Cátia Moreira
  • Roberto Bello-Madruga
  • Marc Torrent Burgas
  • Shina C L Kamerlin
  • Jessica M A Blair
  • Enea Sancho-Vaello

Summary, in English

Efflux is one of the mechanisms employed by Gram-negative bacteria to become resistant to routinely used antibiotics. The inhibition of efflux by targeting their regulators is a promising strategy to re-sensitize bacterial pathogens to antibiotics. AcrAB-TolC is the main resistance-nodulation-division efflux pump in Enterobacteriaceae. MarA is an AraC/XylS family global regulator that regulates more than 40 genes related to the antimicrobial resistance phenotype, including acrAB. The aim of this work was to understand the role of the N-terminal helix of MarA in the mechanism of DNA binding. An N-terminal deletion of MarA showed that the N-terminal helix is critical for recognition of the functional marboxes. By engineering two double cysteine variants of MarA that form a disulfide bond between the N-terminal helix and the hydrophobic core of one of the helices in direct DNA contact, and combining in vitro electrophoretic mobility assays, in vivo measurements of acrAB transcription using a GFP reporter system, and molecular dynamic simulations, it was shown that the immobilization of the N-terminal helix of MarA prevents binding to DNA. This inhibited conformation seems to be universal for the monomeric members of the AraC/XylS family, as suggested by additional molecular dynamics simulations of the two-domain protein Rob. These results point to the N-terminal helix of the AraC/XylS family monomeric regulators as a promising target for the development of inhibitors.

Publishing year

2025-01

Language

English

Publication/Series

Protein Science

Volume

34

Issue

1

Document type

Journal article

Publisher

The Protein Society

Keywords

  • DNA-Binding Proteins/chemistry
  • Molecular Dynamics Simulation
  • Escherichia coli/genetics
  • DNA, Bacterial/metabolism
  • Protein Binding
  • Escherichia coli Proteins/chemistry
  • Protein Domains
  • Bacterial Proteins/chemistry

Status

Published

ISBN/ISSN/Other

  • ISSN: 1469-896X