Structure and Mechanism of RND‐Type Multidrug Efflux Pumps

Nikaido, Hiroshi

doi:10.1002/9780470920541.ch1

Cited by 182 publications

(179 citation statements)

References 139 publications

(266 reference statements)

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“…For example, many of the substrates and inhibitors of AcrB contain weakly acidic or basic groups, and their charge states are likely to become altered in a low dielectric constant environment of the binding site; this is not taken into account in the standard MD simulation procedures currently in use (37). Finally, our system may be incomplete because AcrA appears to be needed for the activity, and thus for the active conformation, of AcrB (6).…”

Section: Discussionmentioning

confidence: 99%

“…However, it was difficult to compare the details of binding of various substrates as determined by MD simulation with their experimentally determined binding affinities. Unfortunately, a simple ligand binding assay (38) does not indicate the affinity to the distal binding pocket, because the binding could have occurred at any other accessible sites on the giant AcrB protein (6). So far, only the export of β-lactams by AcrB has been analyzed in detail (39,40), but K M values are obviously not directly correlated with the K D values of the substrates.…”

Section: Discussionmentioning

confidence: 99%

“…Substrates of these pumps include antibiotics and biocides with very different structural features, although they all seem to have a significant lipophilic portion (5). RND pumps are tripartite complexes (5,6). In the major MDR efflux pump in Escherichia coli, AcrAB-TolC, the RND secondary transporter AcrB, driven by the proton-motive force (7), determines the substrate specificity.…”

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confidence: 99%

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Multidrug binding properties of the AcrB efflux pump characterized by molecular dynamics simulations

Vargiu

Nikaido

2012

Proc. Natl. Acad. Sci. U.S.A.

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Multidrug resistance in Gram-negative bacteria, to which multidrug efflux pumps such as the AcrB transporter makes a major contribution, is becoming a major public health problem. Unfortunately only a few compounds have been cocrystallized with AcrB, and thus computational approaches are essential in elucidating the interaction between diverse ligands and the pump protein. We used molecular dynamics simulation to examine the binding of nine substrates, two inhibitors, and two nonsubstrates to the distal binding pocket of AcrB, identified earlier by X-ray crystallography. This approach gave us more realistic views of the binding than the previously used docking approach, as the explicit water molecules contributed to the process and the flexible binding site was often seen to undergo large structural changes. We analyzed the interaction in detail in terms of the binding energy, hydrophobic surface-matching, and the residues involved in the process. We found that all substrates tested bound to the pocket, whereas the binding to this site was not preferred for the nonsubstrates. Interestingly, both inhibitors [Phe-Arg-β-naphthylamide and 1-(1-naphtylmethyl)-piperazine] tended to move out of the pocket at least partially, getting into contact with a glycine-rich loop that separates the distal pocket from the more proximal region of the protein and is thought to control the access of substrates to the distal pocket.bacterial drug efflux pump | drug-binding pocket | efflux inhibitors

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Multidrug binding properties of the AcrB efflux pump characterized by molecular dynamics simulations

Vargiu

Nikaido

2012

Proc. Natl. Acad. Sci. U.S.A.

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337

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“…The major efflux pump in E. coli is AcrAB-TolC. Although the architecture, assembly, and function of AcrAB-TolC are well known (50,51), and the transcriptional control of the genes encoding AcrAB and TolC have been examined (46,(52)(53)(54)(55)(56), very little has been done to study posttranscriptional regulation. We investigated the role of Hfq-dependent sRNAs in the regulation of genes encoding the efflux pump.…”

Section: Discussionmentioning

confidence: 99%

Small RNA Regulation of TolC, the Outer Membrane Component of Bacterial Multidrug Transporters

Parker

Gottesman

2016

J Bacteriol

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Bacteria use multidrug efflux pumps to export drugs and toxic compounds out of the cell. One of the most important efflux pumps in Escherichia coli is the AcrAB-TolC system. Small regulatory RNAs (sRNAs) are known to be major posttranscriptional regulators that can enhance or repress translation by binding to the 5= untranslated region (UTR) of mRNA targets with the help of a chaperone protein, Hfq. In this study, we investigated the expression of acrA, acrB, and tolC translational fusions using 27 Hfq-dependent sRNAs overexpressed from plasmids. No significant sRNA regulation of acrA or acrB was detected. SdsR (also known as RyeB), an abundant and well-conserved stationary-phase sRNA, was found to repress the expression of tolC, the gene encoding the outer membrane protein of many multidrug resistance efflux pumps. This repression was shown to be by direct base pairing occurring upstream from the ribosomal binding site. SdsR overexpression and its regulation of tolC were found to reduce resistance to novobiocin and crystal violet. Our results suggest that additional targets for SdsR exist that contribute to increased antibiotic sensitivity and reduced biofilm formation. In an effort to identify phenotypes associated with single-copy SdsR and its regulation of tolC, the effect of a deletion of sdsR or mutations in tolC that should block SdsR pairing were investigated using a Biolog phenotypic microarray. However, no significant phenotypes were identified. Therefore, SdsR appears to modulate rather than act as a major regulator of its targets. IMPORTANCEAcrAB-TolC is a major efflux pump present in E. coli and Gram-negative bacteria used to export toxic compounds; the pump confers resistance to many antibiotics of unrelated classes. In this study, we found that SdsR, a small RNA expressed in stationary phase, repressed the expression of tolC, resulting in increased sensitivity to some antibiotics. This extends the findings of previous studies showing that sRNAs contribute to the regulation of many outer membrane proteins; manipulating or enhancing their action might help in sensitizing bacteria to antibiotics.

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“…Furthermore, the observation (4,14) that for the efficient efflux of drugs from cytosol, tripartite pumps, such as AcrB-AcrA-TolC, require help from singlet efflux pumps that extrude drugs into the periplasm suggests that the cytosolic capture by AcrB is at least inefficient. Although proteoliposome reconstitution of an AcrB homolog, AcrD, seemed to suggest that aminoglycosides are captured both from the periplasm and cytosol (1), streptomycin showed no evidence of cytosolic capture, and a recent reexamination of the data suggested that cytosolic capture of other aminoglycosides is marginal at best (8). However, the recent demonstration that the AcrB homolog MexB was active when expressed in a Gram-positive host (15) suggests that such capture does occur.…”

mentioning

confidence: 99%

Vestibules Are Part of the Substrate Path in the Multidrug Efflux Transporter AcrB of Escherichia coli

Husain¹,

Bikhchandani²,

Nikaido³

2011

Journal of Bacteriology

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Structure and Mechanism of RND‐Type Multidrug Efflux Pumps

Cited by 182 publications

References 139 publications

Multidrug binding properties of the AcrB efflux pump characterized by molecular dynamics simulations

Multidrug binding properties of the AcrB efflux pump characterized by molecular dynamics simulations

Small RNA Regulation of TolC, the Outer Membrane Component of Bacterial Multidrug Transporters

Vestibules Are Part of the Substrate Path in the Multidrug Efflux Transporter AcrB of Escherichia coli

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