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Fragment-based screening identifies novel targets for inhibitors of conjugative transfer of antimicrobial resistance by plasmid pKM101.

ABSTRACT: The increasing frequency of antimicrobial resistance is a problem of global importance. Novel strategies are urgently needed to understand and inhibit antimicrobial resistance gene transmission that is mechanistically related to bacterial virulence functions. The conjugative transfer of plasmids by type IV secretion systems is a major contributor to antimicrobial resistance gene transfer. Here, we present a structure-based strategy to identify inhibitors of type IV secretion system-mediated bacterial conjugation. Using differential scanning fluorimetry we screened a fragment library and identified molecules that bind the essential TraE protein of the plasmid pKM101 conjugation machinery. Co-crystallization revealed that fragments bind two alternative sites of the protein and one of them is a novel inhibitor binding site. Based on the structural information on fragment binding we designed novel small molecules that have improved binding affinity. These molecules inhibit the dimerization of TraE, bind to both inhibitor binding sites on TraE and inhibit the conjugative transfer of plasmid pKM101. The strategy presented here is generally applicable for the structure-based design of inhibitors of antimicrobial resistance gene transfer and of bacterial virulence.

SUBMITTER: Casu B

PROVIDER: S-EPMC5668240 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Fragment-based screening identifies novel targets for inhibitors of conjugative transfer of antimicrobial resistance by plasmid pKM101.

Casu Bastien B Arya Tarun T Bessette Benoit B Baron Christian C

Scientific reports 20171102 1

The increasing frequency of antimicrobial resistance is a problem of global importance. Novel strategies are urgently needed to understand and inhibit antimicrobial resistance gene transmission that is mechanistically related to bacterial virulence functions. The conjugative transfer of plasmids by type IV secretion systems is a major contributor to antimicrobial resistance gene transfer. Here, we present a structure-based strategy to identify inhibitors of type IV secretion system-mediated bact ...[more]

PMID: 29097752

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Project description:Multiwalled carbon nanotubes (MWCNTs) regularly enter aquatic environments due to their ubiquity in consumer products and engineering applications. However, the effects of MWCNT pollution on the environmental microbiome are poorly understood. Here, we evaluated whether these carbon nanoparticles can elevate the spread of antimicrobial resistance by promoting bacterial plasmid transfer, which has previously been observed for copper nanomaterials with antimicrobial properties as well as for microplastics. Through a combination of experimental liquid mating assays between Pseudomonas putida donor and recipient strains with plasmid pKJK5::gfpmut3b and mathematical modeling, we here demonstrate that the presence of MWCNTs leads to increased plasmid transfer rates in a concentration-dependent manner. The percentage of transconjugants per recipient significantly increased from 0.21 ± 0.04% in absence to 0.41 ± 0.09% at 10 mg L-1 MWCNTs. Similar trends were observed when using an Escherichia coli donor hosting plasmid pB10. The identified mechanism underlying the observed dynamics was the agglomeration of MWCNTs. A significantly increased number of particles with >6 μm diameter was detected in the presence of MWCNTs, which can in turn provide novel surfaces for bacterial interactions between donor and recipient cells after colonization. Fluorescence microscopy confirmed that MWCNT agglomerates were indeed covered in biofilms that contained donor bacteria as well as elevated numbers of green fluorescent transconjugant cells containing the plasmid. Consequently, MWCNTs provide bacteria with novel surfaces for intense cell-to-cell interactions in biofilms and can promote bacterial plasmid transfer, hence potentially elevating the spread of antimicrobial resistance. IMPORTANCE In recent decades, the use of carbon nanoparticles, especially multiwalled carbon nanotubes (MWCNTs), in a variety of products and engineering applications has been growing exponentially. As a result, MWCNT pollution into environmental compartments has been increasing. We here demonstrate that the exposure to MWCNTs can affect bacterial plasmid transfer rates in aquatic environments, an important process connected to the spread of antimicrobial resistance genes in microbial communities. This is mechanistically explained by the ability of MWCNTs to form bigger agglomerates, hence providing novel surfaces for bacterial interactions. Consequently, increasing pollution with MWCNTs has the potential to elevate the ongoing spread of antimicrobial resistance, a major threat to human health in the 21st century.

Dataset Information

Fragment-based screening identifies novel targets for inhibitors of conjugative transfer of antimicrobial resistance by plasmid pKM101.

Publications

Fragment-based screening identifies novel targets for inhibitors of conjugative transfer of antimicrobial resistance by plasmid pKM101.

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