Multidrug resistance mediated by a bacterial homolog of the human multidrug transporter MDR1.

Veen, Hendrik W. van; Venema, Koen; Bolhuis, Henk; Oussenko, Irina; Kok, Jan; Poolman, Bert; Driessen, Arnold J. M.; Konings, Wil N.

doi:10.1073/pnas.93.20.10668

Cited by 266 publications

(219 citation statements)

References 34 publications

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“…Fluorimetric Analysis of QacA-and QacB-mediated Efflux of DAPI, DiOC 3 , Ethidium, and Pyronin Y-Fluorimetric assays of QacA-and QacB-mediated efflux of DAPI, DiOC 3 , ethidium, and pyronin Y were carried out essentially as described previously for ethidium (8). These substrates bind to specific cellular components and in so doing undergo a change in fluorescence that can be used to indicate how much of the substrate is retained by the cell, e.g.…”

Section: Methodsmentioning

confidence: 99%

“…Currently, all known ATP-dependent multidrug efflux systems belong to the ABC superfamily of transporters, and the best characterized of these is human P-glycoprotein, which confers resistance to an extensive array of cytotoxic, chemotherapeutic agents used in human cancer cells (1,2). Another example is LmrA, a P-glycoprotein homolog that has been identified in Lactococcus lactis and represents the first ATP-dependent multidrug efflux system found in bacteria (3). PMF-dependent multidrug efflux proteins have been classified into three distinct families of membrane transport proteins based on comparative sequence analysis: the major facilitator superfamily (4 -6), e.g.…”

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confidence: 99%

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Bioenergetics of the Staphylococcal Multidrug Export Protein QacA

Mitchell¹,

Paulsen²,

Brown

et al. 1999

Journal of Biological Chemistry

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The multidrug efflux pump QacA from Staphylococcus aureus confers resistance to an extensive range of structurally dissimilar compounds. Fluorimetric analyses demonstrated that QacA confers resistance to the divalent cation 4,6-diamidino-2-phenylindole, utilizing a proton motive force-dependent efflux mechanism previously demonstrated for QacA-mediated resistance to the monovalent cation ethidium. Both the ionophores nigericin and valinomycin inhibited QacA-mediated export of ethidium, indicating an electrogenic drug/nH ؉ (n > 2) antiport mechanism. The kinetic parameters, K m and V max , were determined for QacA-mediated export of four fluorescent substrates, 4,6-diamidino-2-phenylindole, 3,3-dipropyloxacarbocyanine, ethidium, and pyronin Y. Competition studies showed that QacA-mediated ethidium export is competitively inhibited by monovalent cations, e.g. benzalkonium, and non-competitively inhibited by divalent cations, e.g. propamidine, which suggests that monovalent and divalent cations bind at distinct sites on the QacA protein. The quaternary ammonium salt, 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene, was used as a membrane-specific fluorescence probe and demonstrated that the amount of substrate entering the inner leaflet was significantly reduced in QacA-containing strains, supporting the notion that the substrate is extruded directly from the membrane.

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Bioenergetics of the Staphylococcal Multidrug Export Protein QacA

Mitchell¹,

Paulsen²,

Brown

et al. 1999

Journal of Biological Chemistry

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“…5). LmrA belongs to a superfamily of transporters that contain an ATP binding cassette (ABC) 1 (6). It shares significant sequence similarity with members of the P-glycoprotein subfamily of ABC transporters, most notably the human multidrug resistance P-glycoprotein (6).…”

mentioning

confidence: 99%

“…LmrA belongs to a superfamily of transporters that contain an ATP binding cassette (ABC) 1 (6). It shares significant sequence similarity with members of the P-glycoprotein subfamily of ABC transporters, most notably the human multidrug resistance P-glycoprotein (6). This human P-glycoprotein, which is also called MDR1, plays a crucial role in the resistance of cancer cells against cytotoxic agents used in chemotherapy (2).…”

mentioning

confidence: 99%

The Transmembrane Domains of the ABC Multidrug Transporter LmrA Form a Cytoplasmic Exposed, Aqueous Chamber within the Membrane

Poelarends¹,

Konings

2002

Journal of Biological Chemistry

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The ABC multidrug transporter LmrA of Lactococcus lactis consists of six putative transmembrane segments (TMS) and a nucleotide binding domain. LmrA functions as a homodimer in which the two membrane domains form the solute translocation path across the membrane. To obtain structural information of LmrA a cysteine scanning accessibility approach was used. Cysteines were introduced in the cysteine-less wild-type LmrA in each hydrophilic loop and in TMS 6, and each membrane-embedded aromatic residue was mutated to cysteine. Of the 41 constructed single cysteine mutants, only one mutant, L301C, was not expressed. Most singlecysteine mutants were capable of drug transport and only three mutants, F37C, M299C, and N300C, were inactive, indicating that none of the aromatic residues in the transmembrane regions of LmrA are crucial for substrate binding or transport. Modification of the active mutants with N-ethylmaleimide blocked the transport activity in five mutants (S132C, L174C, S206C, S234C, and L292C). All cysteine residues in external and internal loops were accessible to fluorescein maleimide. The labeling experiments also showed that this thiol reagent cannot cross the membrane under the conditions used and confirmed the presence of six TMSs in each monomeric half of the transporter. Surprisingly, several single cysteines in the predicted TMSs could also be labeled by the bulky fluorescein maleimide molecule, suggesting unrestricted accessibility via an aqueous pathway. The periodicity of fluorescein maleimide accessibility of residues 291 to 308 in TMS 6 showed that this membrane-spanning ␣-helix has one face of the helix exposed to an aqueous cavity along its full-length. This finding, together with the solvent accessibility of 11 of 15 membrane-embedded aromatic residues, indicates that the transmembrane domains of the LmrA transporter form, under nonenergized conditions, an aqueous chamber within the membrane, which is open to the intracellular milieu.

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“…This efflux pump belongs to the ATP-binding cassette (ABC) superfamily of transporters and consists of two halves, each half including a hydrophobic transmembrane region or transmembrane domain with six putative a helices and a nucleotide-binding domain; these two halves cooperate to form a single full transporter. Through ATP hydrolysis, this protein has the ability to extrude several amphiphilic compounds from cells against a substrate concentration gradient (van Veen et al, 1996(van Veen et al, , 2006. Homologues to MDR1 are found almost exclusively in eukaryotes, although they have also been found and characterized in some prokaryotes, e.g.…”

Section: Introductionmentioning

confidence: 99%

Identification of a novel ATP-binding cassette transporter involved in long-chain fatty acid import and its role in triacylglycerol accumulation in Rhodococcus jostii RHA1

Villalba

Alvarez

2014

Microbiology

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Members of the genus Rhodococcus are specialists in the biosynthesis and accumulation of triacylglycerols (TAGs). As no transport protein related to TAG metabolism has yet been characterized in these bacteria, we used the available genomic information of Rhodococcus jostii RHA1 to perform a broad survey of genes coding for putative lipid transporter proteins in this oleaginous micro-organism. Among the seven genes encoding putative lipid transporters, ro05645 (now called ltp1: lipid transporter protein) coding for an ATP-binding cassette protein was found clustered with others genes encoding enzymes catalysing the three putative acylation reactions of the Kennedy pathway for TAG synthesis. Overexpression of ltp1 in the RHA1 strain led to an increase of approximately sixfold and threefold in biomass and TAG production, respectively, when cells were cultivated on palmitic acid and oleic acid. Moreover, overexpression of ltp1 also promoted a significant increase in the uptake of a fluorescently labelled long-chain fatty acid (LCFA), as compared with the WT strain RHA1, and its further incorporation into the TAG fraction. Gluconate-grown cells showed increasing amounts of intracellular free fatty acids, but not of TAG, after overexpressing ltp1. Thus, for the first time to our knowledge, a transporter functionally related to TAG metabolism was identified in oleaginous rhodococci. Our results suggested that Ltp1 is an importer of LCFAs that plays a functional role in lipid homeostasis of R. jostii RHA1.

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Multidrug resistance mediated by a bacterial homolog of the human multidrug transporter MDR1.

Cited by 266 publications

References 34 publications

Bioenergetics of the Staphylococcal Multidrug Export Protein QacA

Bioenergetics of the Staphylococcal Multidrug Export Protein QacA

The Transmembrane Domains of the ABC Multidrug Transporter LmrA Form a Cytoplasmic Exposed, Aqueous Chamber within the Membrane

Identification of a novel ATP-binding cassette transporter involved in long-chain fatty acid import and its role in triacylglycerol accumulation in Rhodococcus jostii RHA1

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