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)

Data:	2024
Resum:	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.
Ajuts:	European Commission. Horizon 2020 839036 European Commission. Horizon 2020 890562 Agència de Gestió d'Ajuts Universitaris i de Recerca PID2020-114627RB-I00
Drets:	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.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	AraC/XylS family ; Efflux pump regulation ; MarA ; Mechanism of inhibition ; Resistance-nodulation-division (RND) superfamily ; Rob
Publicat a:	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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