Abstract
Antimicrobial resistance (AMR) in pathogenic microorganisms with multidrug resistance (MDR) constitutes a severe threat to human health. A major causative mechanism of AMR is mediated through the multidrug efflux pump (MEP). The resistance-nodulation-division superfamily (RND family) of Gram-negative bacteria is usually the major cause of MDR in clinical studies. In Salmonella enterica, the RND pump is translated from the acrAB gene, which is regulated by the activator RamA. Many MEP-caused AMR strains have high ramA gene expression due to mutations in RamR, which has a homodimeric structure comprising the dimerization domain and DNA-binding domain (DBD). Three mutations on the dimerization domain, namely Y59H, M84I, and E160D, are far from the DBD; the molecular mechanism through which they influence RamR’s binding affinity to the ramA gene promoter and consequently disrupt RamA remains unclear. The present study conducted molecular dynamics simulations, binding free energy calculations, and normal mode analysis to investigate the mechanism through which Y59H, M84I, and E160D mutations on the dimerization domain influence the binding affinity of RamR to the ramA promoter. The present results suggest that the three mutations alter the RamR structure, resulting in decreased DNA-binding affinity.
Highlights
Antimicrobial resistance (AMR) in pathogenic microorganisms with multidrug resistance (MDR) constitutes a severe threat to human health
Y59 is located at the boundary of the dimerization domain and the DNA-binding domain (DBD); M84 is at the top of the dimerization domain; and E160 is at the dimerization surface between the two monomers
The present study reveals that Y59H, M84I, and E160D mutations in the dimerization domain of RamR, which have been reported as MDR-relevant substitutions[22,25,26], might affect RamR’s binding affinity to the ramA promoter by altering the dynamics properties of the DBD
Summary
Antimicrobial resistance (AMR) in pathogenic microorganisms with multidrug resistance (MDR) constitutes a severe threat to human health. Many MEP-caused AMR strains have high ramA gene expression due to mutations in RamR, which has a homodimeric structure comprising the dimerization domain and DNA-binding domain (DBD). The present study conducted molecular dynamics simulations, binding free energy calculations, and normal mode analysis to investigate the mechanism through which Y59H, M84I, and E160D mutations on the dimerization domain influence the binding affinity of RamR to the ramA promoter. Several studies on RND family-caused bacterial MDR17,18 have reported that RamA, the activator of the acrAB gene, is an essential target for elucidating the MDR mechanism, in S. enterica[19] and K. pneumonia[20,21]. The present study conducted molecular dynamics (MD) simulations and binding free energy calculations to investigate the mechanism through which Y59H, M84I, and E160D mutations in the dimerization domain cause MDR by assessing the binding events between the RamR protein and the ramA promoter. The binding free energy calculations and NMA results were in accordance with the observations from MD simulations
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