Targeting N-terminal domain dynamics to prevent binding
Corbella, Marina (Universitat de Barcelona)
Moreira, Cátia (Uppsala University)
Bello Madruga, Roberto (Universitat Autònoma de Barcelona)
Torrent Burgas, Marc (Universitat Autònoma de Barcelona)
Kamerlin, Shina C. L. (Georgia Institute of Technology)
Blair, Jessica M. A. (University of Birmingham (Regne Unit))
Sancho Vaello, Enea (Universitat Autònoma de Barcelona)

Date:	2024
Abstract:	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.
Grants:	European Commission 839036 European Commission 890562 Agencia Estatal de Investigación PID2020-114627RB-I00
Rights:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Language:	Anglès
Document:	Article ; recerca ; Versió publicada
Subject:	AraC/XylS family ; Efflux pump regulation ; MarA ; Mechanism of inhibition ; Resistance-nodulation-division (RND) superfamily ; Rob
Published in:	Protein science, Vol. 34 (december 2024) , ISSN 1469-896X

DOI: 10.1002/pro.5258
PMID: 39660948

21 p, 32.8 MB

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