Discovery of small molecule inhibitors of xyloglucan endotransglucosylase (XET) activity by high-throughput screening

Chormova, Dimitra; Franková, Lenka; Defries, Andrew; Cutler, Sean R.; Fry, Stephen C.

doi:10.1016/j.phytochem.2015.06.016

Cited by 13 publications

(14 citation statements)

References 55 publications

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“…1 O 2 is produced by riboflavin (in the presence of air containing ordinary oxygen, 3 O 2 ) in the light but not in the dark [ 40 , 70 , 85 ], so samples incubated with riboflavin in the dark or without riboflavin in the light serve as ROS-free controls. 1 O 2 production from 3 O 2 by riboflavin in the light is not stoichiometric: each riboflavin molecule can generate many 1 O 2 molecules, although the yield is not unlimited because riboflavin itself is subject to oxidation by 1 O 2 [ 70 ].…”

Section: Resultsmentioning

confidence: 99%

“…in the presence of tryptophan; [ 37 , 38 ]) and visible light (e.g. in the presence of riboflavin; [ 39 , 40 ]), and has been reported to be responsible for the majority of photo-oxidative damage in leaves [ 41 ]. • OH is the most potent but also the shortest-lived ROS [ 42 , 43 ].…”

Section: Introductionmentioning

confidence: 99%

“…On the other hand, excess ROS can damage cells and are widely implicated in ageing and diseases such as atherosclerosis [ 61 , 62 ]. For example, superoxide can attack membranes, especially their unsaturated fatty acid residues; • OH can react with DNA, proteins, and lipids, causing mutation, denaturation, and membrane permeabilisation, respectively; and 1 O 2 can inactivate enzymes, including xyloglucan endotransglucosylase/hydrolases [ 40 ]. Quenching of ROS may therefore often be beneficial and is potentially achieved by AA, DHA, and DKG.…”

Section: Introductionmentioning

confidence: 99%

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The oxidation of dehydroascorbic acid and 2,3-diketogulonate by distinct reactive oxygen species

Dewhirst

Fry

2018

Biochemical Journal

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l-Ascorbate, dehydro-l-ascorbic acid (DHA), and 2,3-diketo-l-gulonate (DKG) can all quench reactive oxygen species (ROS) in plants and animals. The vitamin C oxidation products thereby formed are investigated here. DHA and DKG were incubated aerobically at pH 4.7 with peroxide (H2O2), ‘superoxide’ (a ∼50 : 50 mixture of and ), hydroxyl radicals (•OH, formed in Fenton mixtures), and illuminated riboflavin (generating singlet oxygen, 1O2). Products were monitored electrophoretically. DHA quenched H2O2 far more effectively than superoxide, but the main products in both cases were 4-O-oxalyl-l-threonate (4-OxT) and smaller amounts of 3-OxT and OxA + threonate. H2O2, but not superoxide, also yielded cyclic-OxT. Dilute Fenton mixture almost completely oxidised a 50-fold excess of DHA, indicating that it generated oxidant(s) greatly exceeding the theoretical •OH yield; it yielded oxalate, threonate, and OxT. 1O2 had no effect on DHA. DKG was oxidatively decarboxylated by H2O2, Fenton mixture, and 1O2, forming a newly characterised product, 2-oxo-l-threo-pentonate (OTP; ‘2-keto-l-xylonate’). Superoxide yielded negligible OTP. Prolonged H2O2 treatment oxidatively decarboxylated OTP to threonate. Oxidation of DKG by H2O2, Fenton mixture, or 1O2 also gave traces of 4-OxT but no detectable 3-OxT or cyclic-OxT. In conclusion, DHA and DKG yield different oxidation products when attacked by different ROS. DHA is more readily oxidised by H2O2 and superoxide; DKG more readily by 1O2. The diverse products are potential signals, enabling organisms to respond appropriately to diverse stresses. Also, the reaction-product ‘fingerprints’ are analytically useful, indicating which ROS are acting in vivo.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The oxidation of dehydroascorbic acid and 2,3-diketogulonate by distinct reactive oxygen species

Dewhirst

Fry

2018

Biochemical Journal

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“…Recently, a screen for inhibitors of xyloglucan endotransglucosylation reported the reduction of parsley XET activity by BB-R250 [19]. We found that BB-R250 also reduced the XET activity in enzyme extracts prepared from host-invading haustoria of C .…”

Section: Resultsmentioning

confidence: 71%

Activity of xyloglucan endotransglucosylases/hydrolases suggests a role during host invasion by the parasitic plant Cuscuta reflexa

Olsen

Krause

2017

PLoS ONE

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The parasitic vines of the genus Cuscuta form haustoria that grow into other plants and connect with their vascular system, thus allowing the parasite to feed on its host. A major obstacle that meets the infection organ as it penetrates the host tissue is the rigid plant cell wall. In the present study, we examined the activity of xyloglucan endotransglucosylases/hydrolases (XTHs) during the host-invasive growth of the haustorium. The level of xyloglucan endotransglucosylation (XET) activity was found to peak at the penetrating stage of Cuscuta reflexa on its host Pelargonium zonale. In vivo colocalization of XET activity and donor substrate demonstrated XET activity at the border between host and parasite. A test for secretion of XET-active enzymes from haustoria of C. reflexa corroborated this and further indicated that the xyloglucan-modifying enzymes originated from the parasite. A known inhibitor of XET, Coomassie Brilliant Blue R250, was shown to reduce the level of XET in penetrating haustoria of C. reflexa. Moreover, the coating of P. zonale petioles with the inhibitor compound lowered the number of successful haustorial invasions of this otherwise compatible host plant. The presented data indicate that the activity of Cuscuta XTHs at the host-parasite interface is essential to penetration of host plant tissue.

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“…Numerous phenotypic screens for small molecule inhibitors of cell wall biosynthesis have been conducted, and have identified a variety of cellulose biosynthesis inhibitors [ 34 – 37 ]. Small molecule inhibitors of xyloglucan endo-transglycosylase have also been identified [ 38 ]. However, these molecules are structurally diverse, precluding the facile identification of a direct target.…”

Section: Introductionmentioning

confidence: 99%

2-Fluoro-L-Fucose Is a Metabolically Incorporated Inhibitor of Plant Cell Wall Polysaccharide Fucosylation

Villalobos

Wallace

2015

PLoS ONE

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The monosaccharide L-fucose (L-Fuc) is a common component of plant cell wall polysaccharides and other plant glycans, including the hemicellulose xyloglucan, pectic rhamnogalacturonan-I (RG-I) and rhamnogalacturonan-II (RG-II), arabinogalactan proteins, and N-linked glycans. Mutations compromising the biosynthesis of many plant cell wall polysaccharides are lethal, and as a result, small molecule inhibitors of plant cell wall polysaccharide biosynthesis have been developed because these molecules can be applied at defined concentrations and developmental stages. In this study, we characterize novel small molecule inhibitors of plant fucosylation. 2-fluoro-L-fucose (2F-Fuc) analogs caused severe growth phenotypes when applied to Arabidopsis seedlings, including reduced root growth and altered root morphology. These phenotypic defects were dependent upon the L-Fuc salvage pathway enzyme L-Fucose Kinase/ GDP-L-Fucose Pyrophosphorylase (FKGP), suggesting that 2F-Fuc is metabolically converted to the sugar nucleotide GDP-2F-Fuc, which serves as the active inhibitory molecule. The L-Fuc content of cell wall matrix polysaccharides was reduced in plants treated with 2F-Fuc, suggesting that this molecule inhibits the incorporation of L-Fuc into these polysaccharides. Additionally, phenotypic defects induced by 2F-Fuc treatment could be partially relieved by the exogenous application of boric acid, suggesting that 2F-Fuc inhibits RG-II biosynthesis. Overall, the results presented here suggest that 2F-Fuc is a metabolically incorporated inhibitor of plant cellular fucosylation events, and potentially suggest that other 2-fluorinated monosaccharides could serve as useful chemical probes for the inhibition of cell wall polysaccharide biosynthesis.

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Discovery of small molecule inhibitors of xyloglucan endotransglucosylase (XET) activity by high-throughput screening

Cited by 13 publications

References 55 publications

The oxidation of dehydroascorbic acid and 2,3-diketogulonate by distinct reactive oxygen species

The oxidation of dehydroascorbic acid and 2,3-diketogulonate by distinct reactive oxygen species

Activity of xyloglucan endotransglucosylases/hydrolases suggests a role during host invasion by the parasitic plant Cuscuta reflexa

2-Fluoro-L-Fucose Is a Metabolically Incorporated Inhibitor of Plant Cell Wall Polysaccharide Fucosylation

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