Dataset Information

Quantifying the Evolutionary Conservation of Genes Encoding Multidrug Efflux Pumps in the ESKAPE Pathogens To Identify Antimicrobial Drug Targets.

ABSTRACT: Increasing rates of antibiotic-resistant bacterial infection are one of the most pressing contemporary global health concerns. The ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) have been identified as the leading global cause of multidrug-resistant bacterial infections, and overexpression of multidrug efflux (MEX) transport systems has been identified as one of the most critical mechanisms facilitating the evolution of multidrug resistance in ESKAPE pathogens. Despite efforts to develop efflux pump inhibitors to combat antibiotic resistance, the need persists to identify additional targets for future investigations. We evaluated evolutionary pressures on 110 MEX-encoding genes from all annotated ESKAPE organism genomes. We identify several MEX genes under stabilizing selection-representing targets which can facilitate broad-spectrum treatments with evolutionary constraints limiting the potential emergence of escape mutants. We also examine MEX systems being evaluated as drug targets, demonstrating that divergent selection may underlie some of the problems encountered in the development of effective treatments-specifically in relation to the NorA system in S. aureus. This study provides a comprehensive evolutionary context to efflux in the ESKAPE pathogens, which will provide critical context to the evaluation of efflux systems as antibiotic targets. IMPORTANCE Increasing rates of antibiotic-resistant bacterial infection are one of the most pressing contemporary global health concerns. The ESKAPE pathogen group represents the leading cause of these infections, and upregulation of efflux pump expression is a significant mechanism of resistance in these pathogens. This has resulted in substantial interest in the development of efflux pump inhibitors to combat antibiotic-resistant infections; however, no widespread treatments have been developed to date. Our study evaluates an often-underappreciated aspect of resistance-the impact of evolutionary selection. We evaluate selection on all annotated efflux genes in all sequenced ESKAPE pathogens, providing critical context for and insight into current and future development of efflux-targeting treatments for resistant bacterial infections.

SUBMITTER: Brooks LE

PROVIDER: S-EPMC5904435 | biostudies-other | 2018 May-Jun

REPOSITORIES: biostudies-other

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Quantifying the Evolutionary Conservation of Genes Encoding Multidrug Efflux Pumps in the ESKAPE Pathogens To Identify Antimicrobial Drug Targets.

Brooks Lauren E LE Ul-Hasan Sabah S Chan Benjamin K BK Sistrom Mark J MJ

mSystems 20180417 3

Increasing rates of antibiotic-resistant bacterial infection are one of the most pressing contemporary global health concerns. The ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and <i>Enterobacter</i> species) have been identified as the leading global cause of multidrug-resistant bacterial infections, and overexpression of multidrug efflux (MEX) transport systems has been identified as one of the most criti ...[more]

PMID: 29719870

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Project description:RND (Resistance-Nodulation-Division) family transporters are widespread especially among Gram-negative bacteria, and catalyze the active efflux of many antibiotics and chemotherapeutic agents. They have very large periplasmic domains, and form tripartite complexes with outer membrane channels and periplasmic adaptor proteins. AcrAB-TolC complex of Escherichia coli, which pumps out a very wide range of drugs, has been studied most intensively. Early studies showed that the transporter captures even those substrates that cannot permeate across the cytoplasmic membrane, such as dianionic beta-lactams, suggesting that the capture can occur from the periplasm. It was also suggested that the capture occurs from the cytoplasmic membrane/periplasm interface, because most substrates contain a sizable hydrophobic domain; however, this may simply be a reflection of the nature of the binding site within AcrB. Genetic studies of chimeric transporters showed that much of the substrate specificity is determined by their periplasmic domains. Biochemical studies with intact cells recently led to the determination of the kinetic constants of AcrB for some beta-lactams, and the result confirms the old prediction that AcrB is a rather slow pump. Reconstitution of purified AcrB and its relatives showed that the pump is a drug/proton antiporter, that AcrA strongly stimulates the activity of the pump, and that AcrB seems to have a highest affinity for conjugated bile salts. Structural study with mutants of the network of charged residues in the transmembrane domain showed that protonation here produced a far-reaching conformational change, which was found to be present in one of the protomers in the asymmetric crystal structure of the wild-type AcrB. The functional rotatory hypothesis then predicts that the drug bound in the periplasmic domain is extruded through this conformational change initiated by the protonation of one of the residues in the aforementioned network, an idea that was recently supported by disulfide cross-linking as well as by the behavior of linked AcrB protomers.

Project description:The substantial rise in multidrug-resistant bacterial infections is a current global imperative. Cumulative efforts to characterize antimicrobial resistance in bacteria has demonstrated the spread of six families of multidrug efflux pumps, of which resistance-nodulation-cell division (RND) is the major mechanism of multidrug resistance in Gram-negative bacteria. RND is composed of a tripartite protein assembly and confers resistance to a range of unrelated compounds. In the major enteric pathogen Campylobacter jejuni, the three protein components of RND are posttranslationally modified with N-linked glycans. The direct role of N-linked glycans in C. jejuni and other bacteria has long been elusive. Here, we present the first detailed account of the role of N-linked glycans and the link between N-glycosylation and antimicrobial resistance in C. jejuni We demonstrate the multifunctional role of N-linked glycans in enhancing protein thermostability, stabilizing protein complexes and the promotion of protein-protein interaction, thus mediating antimicrobial resistance via enhancing multidrug efflux pump activity. This affirms that glycosylation is critical for multidrug efflux pump assembly. We present a generalized strategy that could be used to investigate general glycosylation system in Campylobacter genus and a potential target to develop antimicrobials against multidrug-resistant pathogens.IMPORTANCE Nearly all bacterial species have at least a single glycosylation system, but the direct effects of these posttranslational protein modifications are unresolved. Glycoproteome-wide analysis of several bacterial pathogens has revealed general glycan modifications of virulence factors and protein assemblies. Using Campylobacter jejuni as a model organism, we have studied the role of general N-linked glycans in the multidrug efflux pump commonly found in Gram-negative bacteria. We show, for the first time, the direct link between N-linked glycans and multidrug efflux pump activity. At the protein level, we demonstrate that N-linked glycans play a role in enhancing protein thermostability and mediating the assembly of the multidrug efflux pump to promote antimicrobial resistance, highlighting the importance of this posttranslational modification in bacterial physiology. Similar roles for glycans are expected to be found in other Gram-negative pathogens that possess general protein glycosylation systems.

Dataset Information

Quantifying the Evolutionary Conservation of Genes Encoding Multidrug Efflux Pumps in the ESKAPE Pathogens To Identify Antimicrobial Drug Targets.

Publications

Quantifying the Evolutionary Conservation of Genes Encoding Multidrug Efflux Pumps in the ESKAPE Pathogens To Identify Antimicrobial Drug Targets.

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