Microbial fermentation-derived inhibitors of efflux-pump-mediated drug resistance

Lee, May D.; Galazzo, Jorge L.; Staley, Andrew L.; Lee, Julie C.; Warren, Mark S.; Fuernkranz, Hans A.; Chamberland, Suzanne; Lomovskaya, Olga; Miller, George H.

doi:10.1016/s0014-827x(01)01002-3

Cited by 93 publications

(69 citation statements)

References 9 publications

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“…The milbemycins are 16-membered ring macrolides belonging to families that are used as insecticides, acaricides, and anthelminthics (44,60). Some milbemycins have also been identified as chemosensitizers of fungal multidrug efflux mediated by CaCdr1p (33). Enniatin is another circular molecule that has been found to inhibit antifungal drug efflux (25,33).…”

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confidence: 99%

Characterization of Three Classes of Membrane Proteins Involved in Fungal Azole Resistance by Functional Hyperexpression inSaccharomyces cerevisiae

et al. 2007

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The study of eukaryotic membrane proteins has been hampered by a paucity of systems that achieve consistent high-level functional protein expression. We report the use of a modified membrane protein hyperexpression system to characterize three classes of fungal membrane proteins (ABC transporters Pdr5p, CaCdr1p, CaCdr2p, CgCdr1p, CgPdh1p, CkAbc1p, and CneMdr1p, the major facilitator superfamily transporter CaMdr1p, and the cytochrome P450 enzyme CaErg11p) that contribute to the drug resistance phenotypes of five pathogenic fungi and to express human P glycoprotein (HsAbcb1p). The hyperexpression system consists of a set of plasmids that direct the stable integration of a single copy of the expression cassette at the chromosomal PDR5 locus of a modified host Saccharomyces cerevisiae strain, AD⌬. Overexpression of heterologous proteins at levels of up to 29% of plasma membrane protein was achieved. Membrane proteins were expressed with or without green fluorescent protein (GFP), monomeric red fluorescent protein, His, FLAG/His, Cys, or His/Cys tags. Most GFP-tagged proteins tested were correctly trafficked within the cell, and His-tagged proteins could be affinity purified. Kinetic analysis of ABC transporters indicated that the apparent K m value and the V max value of ATPase activities were not significantly affected by the addition of His tags. The efflux properties of seven fungal drug pumps were characterized by their substrate specificities and their unique patterns of inhibition by eight xenobiotics that chemosensitized S. cerevisiae strains overexpressing ABC drug pumps to fluconazole. The modified hyperexpression system has wide application for the study of eukaryotic membrane proteins and could also be used in the pharmaceutical industry for drug screening.The resolution and exploitation of protein structure and function are among the greatest biological challenges in the postgenomic era. These challenges, and their potential dividends, are greatest for membrane proteins, which are notoriously difficult to functionally express and purify in the quantities and forms needed for drug discovery or for high-resolution X-ray crystallography (1, 16). About a quarter of the cellular proteome consists of membrane proteins (5), which often play vital physiological roles: from environmental sensing to energy transduction, from nutrient uptake to drug efflux, and from cellular proliferation to programmed cell death. Membrane proteins are involved in many prominent diseases, including cystic fibrosis (48), type 2 diabetes (49), heart disease (52), and the drug resistance of numerous cancers (57). Hence, they are the targets for many therapies and constitute up to 70% of the drug targets used in medicine today. Membrane proteins also play key roles in drug modification, detoxification, and resistance in a wide variety of prokaryotic and eukaryotic systems (7). A fundamental understanding of cell biology, cell physiology, and cell-drug interactions therefore requires a detailed analysis of membrane protein function. Fu...

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Characterization of Three Classes of Membrane Proteins Involved in Fungal Azole Resistance by Functional Hyperexpression inSaccharomyces cerevisiae

et al. 2007

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“…The ethyl acetate extract was purified by silica gel column chromatography twice and HPLC to yield naphthacemycins A 1 -A 11 (1-11), B 1 -B 4 (12)(13)(14)(15) and C 1 -C 2 (16)(17). Their structures were elucidated by NMR experiments and X-ray analysis, as reported elsewhere.…”

Section: Fermentation and Isolationmentioning

confidence: 99%

“…EA-371α (24), produced by Streptomyces sp., was isolated as an inhibitor of the MexAB-OprM efflux pump, which is involved in intrinsic antibiotic resistance of Pseudomonas aeruginosa. 16 Though its skeleton is 8-oxobenzo [a] naphthacene, it has a 1,3-dihydroxy-10,10-dimethylanthrone unit, similar to the naphthacemycins. Its desulfonated analog, benastatin A (25), is active against Gram-positive bacteria.…”

Section: Biological Activitiesmentioning

confidence: 99%

“…The crude material VI was separated by Pegasil ODS HPLC using 75% MeOH (flow rate, 8 ml min À 1 ), and naphthacemycins A 2 (2, 13.9 mg), B 3 (14, 10.0 mg) and B 4 (15, 30.5 mg) and crude materials VIII (20.2 mg) and IX (61.2 mg) were eluted at 17, 19, 27, 22 and 36 min, respectively. The crude material VIII was applied to Pegasil ODS HPLC using 55% CH 3 CN (flow rate, 8 ml min À 1 ), and the eluted peak of 18 min was collected to yield naphthacemycin C 1 (16,14.2 mg). The crude material IX was further purified by Pegasil ODS HPLC using 55% CH 3 CN (flow rate, 8 ml min À 1 ) to yield naphthacemycins A 3 (3, 21.5 mg) and A 4 (4, 32.8 mg) at the eluate of 21 and 24 min, respectively.…”

Section: Isolationmentioning

confidence: 99%

“…2 We have also reported that cyslabdan enhances imipenem activity against MRSA. 3,4 Our continuous study to find microbial metabolites, like cyslabdan, that circumvent β-lactam resistance of MRSA, led us to discover further novel compounds, the naphthacemycins A 1 -A 11 (1-11), B 1 -B 4 (12)(13)(14)(15) and C 1 -C 2 (16)(17) (Figure 1), reported as KB-3346-5 substances in the patent by our group 5 , isolated from a culture broth of Streptomyces sp. KB-3346-5.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Naphthacemycins, novel circumventors of β-lactam resistance in MRSA, produced by Streptomyces sp. KB-3346-5. I. The taxonomy of the producing strain, and the fermentation, isolation and antibacterial activities

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The Potential of Peptides and Depsipeptides from Terrestrial and Marine Organisms in the Fight against Human Protozoan Diseases

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Microbial fermentation-derived inhibitors of efflux-pump-mediated drug resistance

Cited by 93 publications

References 9 publications

Characterization of Three Classes of Membrane Proteins Involved in Fungal Azole Resistance by Functional Hyperexpression inSaccharomyces cerevisiae

Characterization of Three Classes of Membrane Proteins Involved in Fungal Azole Resistance by Functional Hyperexpression inSaccharomyces cerevisiae

Naphthacemycins, novel circumventors of β-lactam resistance in MRSA, produced by Streptomyces sp. KB-3346-5. I. The taxonomy of the producing strain, and the fermentation, isolation and antibacterial activities

The Potential of Peptides and Depsipeptides from Terrestrial and Marine Organisms in the Fight against Human Protozoan Diseases

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