A 1H NMR study of the specificity of α-l-arabinofuranosidases on natural and unnatural substrates

Borsenberger, Vinciane; Dornez, Emmie; Desrousseaux, Marie‐Laure; Massou, Stéphane; Tenkanen, Maija; Courtin, Christophe M.; Dumon, Claire; O’Donohue, Michael; Fauré, Régis

doi:10.1016/j.bbagen.2014.07.001

Cited by 17 publications

(9 citation statements)

References 27 publications

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“…Carbohydrate-active enzymes families that were enriched through growth on poplar hydrolysate included those that comprise enzymes involved in plant polysaccharide deconstruction. For example, family GH43 includes enzymes that target arabinoxylan ( Borsenberger et al, 2014 ; Mewis et al, 2016 ), family CE1 members were shown to deacetylate polymeric xylans ( Neumuller et al, 2015 ; Mai-Gisondi et al, 2017 ), and family GH5 members include endoxylanases that targets xylans with or without methyl-glucuronic acid side chain ( Gallardo et al, 2010 ), as well as enzymes that target cellulose and mannans ( Aspeborg et al, 2012 ). Notably, enzymes belonging to families GH2 and GH3 were also abundant in the moose rumen microbiome, and predicted to participate in plant cell wall deconstruction ( Svartström et al, 2017 ).…”

Section: Resultsmentioning

confidence: 99%

Comparative Metagenomics of Cellulose- and Poplar Hydrolysate-Degrading Microcosms from Gut Microflora of the Canadian Beaver (Castor canadensis) and North American Moose (Alces americanus) after Long-Term Enrichment

Wong¹,

Wang²,

Couturier³

et al. 2017

Front. Microbiol.

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To identify carbohydrate-active enzymes (CAZymes) that might be particularly relevant for wood fiber processing, we performed a comparative metagenomic analysis of digestive systems from Canadian beaver (Castor canadensis) and North American moose (Alces americanus) following 3 years of enrichment on either microcrystalline cellulose or poplar hydrolysate. In total, 9,386 genes encoding CAZymes and carbohydrate-binding modules (CBMs) were identified, with up to half predicted to originate from Firmicutes, Bacteroidetes, Chloroflexi, and Proteobacteria phyla, and up to 17% from unknown phyla. Both PCA and hierarchical cluster analysis distinguished the annotated glycoside hydrolase (GH) distributions identified herein, from those previously reported for grass-feeding mammals and herbivorous foragers. The CAZyme profile of moose rumen enrichments also differed from a recently reported moose rumen metagenome, most notably by the absence of GH13-appended dockerins. Consistent with substrate-driven convergence, CAZyme profiles from both poplar hydrolysate-fed cultures differed from cellulose-fed cultures, most notably by increased numbers of unique sequences belonging to families GH3, GH5, GH43, GH53, and CE1. Moreover, pairwise comparisons of moose rumen enrichments further revealed higher counts of GH127 and CE15 families in cultures fed with poplar hydrolysate. To expand our scope to lesser known carbohydrate-active proteins, we identified and compared multi-domain proteins comprising both a CBM and domain of unknown function (DUF) as well as proteins with unknown function within the 416 predicted polysaccharide utilization loci (PULs). Interestingly, DUF362, identified in iron–sulfur proteins, was consistently appended to CBM9; on the other hand, proteins with unknown function from PULs shared little identity unless from identical PULs. Overall, this study sheds new light on the lignocellulose degrading capabilities of microbes originating from digestive systems of mammals known for fiber-rich diets, and highlights the value of enrichment to select new CAZymes from metagenome sequences for future biochemical characterization.

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Section: Resultsmentioning

confidence: 99%

Comparative Metagenomics of Cellulose- and Poplar Hydrolysate-Degrading Microcosms from Gut Microflora of the Canadian Beaver (Castor canadensis) and North American Moose (Alces americanus) after Long-Term Enrichment

Wong¹,

Wang²,

Couturier³

et al. 2017

Front. Microbiol.

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“…As such, it is possible that the XOSs had arabinose branches. Previous studies provided evidence that the ␣-1,2-and ␣-1,3-L-arabinofuranosyl residues in the arabinobranched XOSs could be identified by 1 H nuclear magnetic resonance ( 1 H-NMR) spectroscopy (24)(25)(26)(27). 1 H-NMR analyses also suggested the presence of 2-O-linked (␦ 5.28) and 3-O-linked (␦ 5.33) arabinoses in the XynA-hydrolyzed WAX products ( Fig.…”

mentioning

confidence: 91%

Two Distinct α- l -Arabinofuranosidases in Caldicellulosiruptor Species Drive Degradation of Arabinose-Based Polysaccharides

Saleh

Han

Lü

et al. 2017

Appl Environ Microbiol

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Species in the extremely thermophilic genus Caldicellulosiruptor can degrade unpretreated plant biomass through the action of multimodular glycoside hydrolases. To date, most focus with these bacteria has been on hydrolysis of glucans and xylans, while the biodegradation mechanism for arabinose-based polysaccharides remains unclear. Here, putative ␣-L-arabinofuranosidases (AbFs) were identified in Caldicellulosiruptor species by homology to less-thermophilic versions of these enzymes. From this screen, an extracellular XynF was determined to be a key factor in hydrolyzing ␣-1,2-, ␣-1,3-, and ␣-1,5-L-arabinofuranosyl residues of arabinosebased polysaccharides. Combined with a GH11 xylanase (XynA), XynF increased arabinoxylan hydrolysis more than 6-fold compared to the level seen with XynA alone, likely the result of XynF removing arabinofuranosyl side chains to generate linear xylans that were readily degraded. A second AbF, the intracellular AbF51, preferentially cleaved the ␣-1,5-L-arabinofuranosyl glycoside bonds within sugar beet arabinan. ␤-Xylosidases, such as GH39 Xyl39B, facilitated the hydrolysis of arabinofuranosyl residues at the nonreducing terminus of the arabinosebranched xylo-oligosaccharides by AbF51. These results demonstrate the separate but complementary contributions of extracellular XynF and cytosolic AbF51 in processing the bioconversion of arabinose-containing oligosaccharides to fermentable monosaccharides.IMPORTANCE Degradation of hemicellulose, due to its complex chemical structure, presents a major challenge during bioconversion of lignocellulosic biomass to biobased fuels and chemicals. Degradation of arabinose-containing polysaccharides, in particular, can be a key bottleneck in this process. Among Caldicellulosiruptor species, the multimodular arabinofuranosidase XynF is present in only selected members of this genus. This enzyme exhibited high hydrolysis activity, broad specificity, and strong synergism with other hemicellulases acting on arabino-polysaccharides. An intracellular arabinofuranosidase, AbF51, occurs in all Caldicellulosiruptor species and, in conjunction with xylosidases, processes the bioconversion of arabinose-branched oligosaccharides to fermentable monosaccharides. Taken together, the data suggest that plant biomass degradation in Caldicellulosiruptor species involves extracellular XynF that acts synergistically with other hemicellulases to digest arabino-polysaccharides that are subsequently transported and degraded further by intracellular AbF51 to produce shortchain arabino sugars.

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“…Fine substrate recognition was studied for the GH43 enzyme from B. adolescentis using two chromogenic artificial substrates enabling one to demonstrate how the selectivity of this enzyme is a property that often remains unstudied due to the lack of appropriate substrates and readily-accessible methods. In an elegant experiment, enzyme-mediated hydrolysis of 1, 2 and 3 ( Figure 3 ) was monitored by 1 H NMR, performing reactions in standard 5-mm NMR tubes [ 25 ], taking advantage of the clear differences in the chemical shifts of the anomeric signals of different arabinose moieties at different positions.…”

Section: Arabinofuranosidasementioning

confidence: 99%

Uncommon Glycosidases for the Enzymatic Preparation of Glycosides

Trincone

2015

Biomolecules

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Most of the reports in literature dedicated to the use of glycosyl hydrolases for the preparation of glycosides are about gluco- (α- and β-form) and galacto-sidase (β-form), reflecting the high-availability of both anomers of glucosides and of β-galactosides and their wide-ranging applications. Hence, the idea of this review was to analyze the literature focusing on hardly-mentioned natural and engineered glycosyl hydrolases. Their performances in the synthetic mode and natural hydrolytic potential are examined. Both the choice of articles and their discussion are from a biomolecular and a biotechnological perspective of the biocatalytic process, shedding light on new applicative ideas and on the assortment of biomolecular diversity. The hope is to elicit new interest for the development of biocatalysis and to gather attention of biocatalyst practitioners for glycosynthesis.

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A 1H NMR study of the specificity of α-l-arabinofuranosidases on natural and unnatural substrates

Cited by 17 publications

References 27 publications

Comparative Metagenomics of Cellulose- and Poplar Hydrolysate-Degrading Microcosms from Gut Microflora of the Canadian Beaver (Castor canadensis) and North American Moose (Alces americanus) after Long-Term Enrichment

Comparative Metagenomics of Cellulose- and Poplar Hydrolysate-Degrading Microcosms from Gut Microflora of the Canadian Beaver (Castor canadensis) and North American Moose (Alces americanus) after Long-Term Enrichment

Two Distinct α- l -Arabinofuranosidases in Caldicellulosiruptor Species Drive Degradation of Arabinose-Based Polysaccharides

Uncommon Glycosidases for the Enzymatic Preparation of Glycosides

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