The ABCs of Candida albicans Multidrug Transporter Cdr1

Prasad, Rajendra; Banerjee, Atanu; Khandelwal, Nitesh; Dhamgaye, Sanjiveeni

doi:10.1128/ec.00137-15

Cited by 98 publications

(81 citation statements)

References 75 publications

(83 reference statements)

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“…), the deviations occur in Walker B (aspartate or serine instead of glutamate), switch loop (glutamine or serine instead of histidine), and ABC signature (substitution of the central glycine by another amino acid), indicating that they play a dedicated function. Of note, there are asymmetric Type II ABC exporters with an inverse domain topology and different TMD architecture, such as Pdr5 , Cdr1 , and ABCG5/ABCG8 . They contain a degenerate ATP‐binding site as well, but the pattern and functional role of their noncanonical substitutions is different from classical Type I ABC exporters.…”

Section: Discussionmentioning

confidence: 99%

Split tasks of asymmetric nucleotide‐binding sites in the heterodimeric ABC exporter EfrCD

2017

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Many heterodimeric ATP-binding cassette (ABC) exporters evolved asymmetric ATP-binding sites containing a degenerate site incapable of ATP hydrolysis due to noncanonical substitutions in conserved sequence motifs. Recent studies revealed that nucleotide binding to the degenerate site stabilizes contacts between the nucleotide-binding domains (NBDs) of the inward-facing transporter and regulates ATP hydrolysis at the consensus site via allosteric coupling mediated by the D-loops. However, it is unclear whether nucleotide binding to the degenerate site is strictly required for substrate transport. In this study, we examined the functional consequences of a systematic set of mutations introduced at the degenerate and consensus site of the multidrug efflux pump EfrCD of Enterococcus faecalis. Mutating motifs which differ among the two ATP-binding sites (Walker B, switch loop, and ABC signature) or which are involved in interdomain communication (D-loop and Q-loop) led to asymmetric results in the functional assays and were better tolerated at the degenerate site. This highlights the importance of the degenerate site to allosterically regulate the events at the consensus site. Mutating invariant motifs involved in ATP binding and NBD closure (A-loop and Walker A) resulted in equally reduced transport activities, regardless at which ATP-binding site they were introduced. In contrast to previously investigated heterodimeric ABC exporters, mutation of the degenerate site Walker A lysine completely inactivated ATPase activity and substrate transport, indicating that ATP binding to the degenerate site is essential for EfrCD. This study provides novel insights into the split tasks of asymmetric ATP-binding sites of heterodimeric ABC exporters.

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Section: Discussionmentioning

confidence: 99%

Split tasks of asymmetric nucleotide‐binding sites in the heterodimeric ABC exporter EfrCD

2017

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“…Multiple mechanisms are responsible for this, with genetic alterations in ATP-binding cassette (ABC) multidrug transporters playing a major role. Often, multidrug resistance results from the overexpression of these proteins in fungal pathogens and cancers (Konotoylannis and Lewis, 2002;Gottesman et al, 2002;Lage, 2003;Pfaller, 2012;Prasad et al, 2014;Kathawala et al, 2015). It seemed plausible that additional mechanisms for increased multidrug resistance existed and were worth identifying.…”

Section: Introductionmentioning

confidence: 99%

An A666G mutation in transmembrane helix 5 of the yeast multidrug transporter Pdr5 increases drug efflux by enhancing cooperativity between transport sites

Arya

Rahman

Rudrow

et al. 2019

Molecular Microbiology

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Summary Resistance to antimicrobial and chemotherapeutic agents is a significant clinical problem. Overexpression of multidrug efflux pumps often creates broad‐spectrum resistance in cancers and pathogens. We describe a mutation, A666G, in the yeast ABC transporter Pdr5 that shows greater resistance to most of the tested compounds than does an isogenic wild‐type strain. This mutant exhibited enhanced resistance without increasing either the amount of protein in the plasma membrane or the ATPase activity. In fluorescence quenching transport assays with rhodamine 6G in purified plasma membrane vesicles, the initial rates of rhodamine 6G fluorescence quenching of both the wild type and mutant showed a strong dependence on the ATP concentration, but were about twice as high in the latter. Plots of the initial rate of fluorescence quenching versus ATP concentration exhibited strong cooperativity that was further enhanced in the A666G mutant. Resistance to imazalil sulfate was about 3–4x as great in the A666G mutant strain as in the wild type. When this transport substrate was used to inhibit the rhodamine 6G transport, the A666G mutant inhibition curves also showed greater cooperativity than the wild‐type strain. Our results suggest a novel and important mechanism: under selection, Pdr5 mutants can increase drug resistance by improving cooperative interactions between drug transport sites.

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“…The latest version of transportDB [64] http://www.membranetransport.org/transportDB2/overview.html, the most comprehensive listing, gives 341 transporters for S. cerevisiae S288c, and shows that improvements in identification have been less than substantial. In general, the efflux transporters seem to have evolved to remove natural toxins from the yeast's environment [65,66], are highly promiscuous [67,68], and many are known as pleiotropic drug transporters (PDRs) [69].…”

Section: A Large Percentage Of Orphans Even In Well-studied Organismsmentioning

confidence: 99%

Control of metabolite efflux in microbial cell factories: current advances and future prospects

Kell¹

2018

Preprint

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The yield and ease of purification of biotechnological products are typically greatly enhanced if they can be secreted into the external medium. Although not universally appreciated, however, small molecule biotechnological products do not ‘float across’ the phospholipid bilayer portion of biological membranes, and thus they need transporters to assist their passage into the extramembrane and extracellular spaces. Some of these transporters may be reversible, equilibrative transporters that might more normally be used for uptake, while others may have an efflux directionality imposed on them via suitable energy coupling mechanisms. Despite the energetic costs of small molecule synthesis, natural evolution does in fact provide a number of mechanisms by which the secretion of such products can actually enhance fitness. Where available these provide useful starting points, and assaying for such activities is crucial. In particular, in a systems biology approach, we first need to identify such/suitable activities before we can seek to increase them. They may then be improved through promoter engineering, via manipulation of control elements, or by directed evolution of the transporter proteins themselves. Modern methods of synthetic biology provide enormous opportunities for all kinds of efflux transporter engineering; they are just beginning to be realised.

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The ABCs of Candida albicans Multidrug Transporter Cdr1

Cited by 98 publications

References 75 publications

Split tasks of asymmetric nucleotide‐binding sites in the heterodimeric ABC exporter EfrCD

Split tasks of asymmetric nucleotide‐binding sites in the heterodimeric ABC exporter EfrCD

An A666G mutation in transmembrane helix 5 of the yeast multidrug transporter Pdr5 increases drug efflux by enhancing cooperativity between transport sites

Control of metabolite efflux in microbial cell factories: current advances and future prospects

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