Development of binding assays to screen ligands for Plasmodium falciparum thioredoxin and glutathione reductases by ultrafiltration and liquid chromatography/mass spectrometry

Mulabagal, Vanisree; Calderón, Ángela I.

doi:10.1016/j.jchromb.2010.02.030

Cited by 25 publications

(13 citation statements)

References 39 publications

(45 reference statements)

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“…Based on the report of Mulabagal and Calderón in 2010 (Mulabagal & Calderón 2010) regarding curcumin and DMC as PfTrxR ligands, we tested these compounds for their ability to inhibit PfTrxR. Curcumin and DMC displayed good binding affinity for the PfTrxR target enzyme but curcumin displayed less than 50% inhibition of PfTrxR at 10 mM when tested in the functional assay compared to DMC which displayed more than 50% PfTrxR inhibition at 10 mM with an IC 50 value of 2.0 mM.…”

Section: Resultsmentioning

confidence: 99%

Determination of antiplasmodial activity and binding affinity of curcumin and demethoxycurcumin towardsPfTrxR

Munigunti

Gathiaka

Acevedo

et al. 2014

Natural Product Research

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In our study, the inhibitory activity of curcuminoids towards Plasmodium falciparum thioredoxin reductase (PfTrxR) was determined using LC-MS-based functional assay and showed that only demethoxycurcumin (DMC) inhibited PfTrxR (IC50: 2 μM). In silico molecular modelling was used to ascertain and further confirm that the binding affinities of curcumin and DMC are towards the dimer interface of PfTrxR. The in vitro antiplasmodial activities of curcumin and DMC were evaluated and shown to be active against chloroquine (CQ)-sensitive (D6 clone) and moderately active against CQ-resistant (W2 clone) strains of Plasmodium falciparum while no cytotoxicity was observed against Vero cells.

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Section: Resultsmentioning

confidence: 99%

Determination of antiplasmodial activity and binding affinity of curcumin and demethoxycurcumin towardsPfTrxR

Munigunti

Gathiaka

Acevedo

et al. 2014

Natural Product Research

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“…Chemicals and enzymes: Solvents for LC-MS analysis were purchased from Fischer Scientific International (Atlanta, GA). Eight compounds from AnalytiCon Discovery GmbH (Germany) natural products library (MEGx) were purchased based on compound classes that have reported either antimalarial activity or inhibition of TrxR [5,7]. Hispolone was kindly provided by Dr. G. V. Subbaraju, AptuitLauras, Hyderabad, India.…”

Section: Methodsmentioning

confidence: 99%

Determination of Antiplasmodial Activity and Binding Affinity of Selected Natural Products towards PfTrxR and PfGR

Munigunti

Becker

Brun

et al. 2013

Natural Product Communications

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In our study, the binding affinities of selected natural products towards PfTrxR, PfGR, human TrxR and human GR were determined using a mass spectrometry based ligand binding assay. The in vitro antimalarial activity and cytotoxicity of these ligands were also determined. Catharanthine, 11-(OH)-coronaridine, hernagine, vobasine and hispolone displayed antiplasmodial activity against PfK1 (IC 50 = 0.996-3.63 μg/mL).

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“…All the plant extracts were tested for their binding affinity at 100 µg/mL. The binding assay was performed according to the published method [4]. The ligands were reconstituted in 100 µL of MeOH _ H 2 O (80 : 20, v/v) and were analyzed by LC-MS. Denatured enzyme was used for the controlled experiment, and assays were carried out in duplicate.…”

Section: Pftrxr Enzyme Binding Assay Using Ultrafiltration (Uf) and Lmentioning

confidence: 99%

“…Plant extracts contain a large number of different chemical constituents making it difficult to ascertain their antimalarial activity before isolation of the bioactive compounds. A suitable tool to speed up natural product drug discovery is the application of UF/LC-MS-based binding assays [4] to screen complex mixtures derived from plants for their binding affinity towards PfTrxR followed by MS-based structure elucidation and confirmation of the identity of new ligands.…”

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confidence: 99%

Identification of Oleamide inGuatteria recurvisepalaby LC/MS-BasedPlasmodium falciparumThioredoxin Reductase Ligand Binding Method

Munigunti¹,

Nelson²,

Mulabagal³

et al. 2011

Planta Med

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Our current research on applications of mass spectrometry to natural product drug discovery against malaria aims to screen plant extracts for new ligands to Plasmodium falciparum thioredoxin reductase (PfTrxR) followed by their identification and structure elucidation. PfTrxR is involved in the antioxidant defense and redox regulation of the parasite and is validated as a promising target for therapeutic intervention against malaria. In the present study, detannified methanol extracts from Guatteria recurvisepala, Licania kallunkiae, and Topobea watsonii were screened for ligands to PfTrxR using ultrafiltration and liquid chromatography/mass spectrometry-based binding experiments. The PfTrxR ligand identified in the extract of Guatteria recurvisepala displayed a relative binding affinity of 3.5-fold when incubated with 1 μM PfTrxR. The ligand corresponding to the protonated molecule m/z 282.2792 [M+ H]+ was eluted at a retention time of 17.95 min in a 20-min gradient of 95% B consisting of (A) 0.1%formic acid in 95% H₂O-5% ACN, and (B) 0.1% formic acid in 95% ACN-5% H₂O in an LC-QTOF-MS.Tandem MS of the protonated molecule m/z 282.2792 [M + H]+, C₁₈H₃₆NO (DBE: 2; error: 1.13 ppm) resulted in two daughter ions m/z 265.2516[M + H-NH₃]+ (DBE: 3; error: 0.35 ppm) and m/z 247.2405 [M + H-NH₃-H₂O] +, (DBE: 4; error:2.26 ppm). The PfTrxR ligand was identified as oleamide and confirmed by comparison of the retention time, molecular formula, accurate mass,and double bond equivalence with the standard oleamide. This is the first report on the identification of oleamide as a PfTrxR ligand from Guatteria recurvisepala R. E. Fr. and the corresponding in vitro activity against P. falciparum strain K1 (IC₅₀ 4.29 μg/mL).

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Development of binding assays to screen ligands for Plasmodium falciparum thioredoxin and glutathione reductases by ultrafiltration and liquid chromatography/mass spectrometry

Cited by 25 publications

References 39 publications

Determination of antiplasmodial activity and binding affinity of curcumin and demethoxycurcumin towardsPfTrxR

Determination of antiplasmodial activity and binding affinity of curcumin and demethoxycurcumin towardsPfTrxR

Determination of Antiplasmodial Activity and Binding Affinity of Selected Natural Products towards PfTrxR and PfGR

Identification of Oleamide inGuatteria recurvisepalaby LC/MS-BasedPlasmodium falciparumThioredoxin Reductase Ligand Binding Method

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