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(Bifidobacterium adolescentis)

Product code: E-AFAM2

400 Units on WAX at 40oC

Prices exclude VAT

This product has been discontinued

Content: 400 Units on WAX at 40oC
Shipping Temperature: Ambient
Storage Temperature: 2-8oC
Formulation: In 3.2 M ammonium sulphate
Physical Form: Suspension
Stability: > 4 years at 4oC
Enzyme Activity: α-Arabinofuranosidase
EC Number:
CAZy Family: GH43
CAS Number: 9067-74-7
Synonyms: non-reducing end alpha-L-arabinofuranosidase; alpha-L-arabinofuranoside non-reducing end alpha-L-arabinofuranosidase
Source: Bifidobacterium adolescentis
Molecular Weight: 59,404
Concentration: Supplied at ~ 200 U/mL
Expression: Recombinant from Bifidobacterium adolescentis
Specificity: Highly specific hydrolysis of α-1,3-linked L-arabinofuranose residues from doubly substituted D-xylosyl or L-arabinosyl residues of arabinoxylans and branched arabinans, respectively.
Specific Activity: ~ 90 U/mg (40oC, pH 6.0 on xylanase degraded wheat arabinoxylan);
~ 28 U/mg (40oC, pH 6.0 on wheat arabinoxylan);
~ 4 U/mg (40oC, pH 6.0 on sugar-beet arabinan);
~ 0.1 U/mg (40oC, pH 6.0 on p-nitrophenyl-α-L-arabinofuranoside)
Unit Definition: One Unit of α-L-arabinofuranosidase activity is defined as the amount of enzyme required to release one µmole of arabinose per minute from wheat arabinoxylan (10 mg/mL) in sodium phosphate buffer (100 mM), pH 6.0 at 40oC.
Temperature Optima: 50oC
pH Optima: 6
Application examples: Applications in carbohydrate and biofuels research.

This product has been discontinued (read more).

High purity α-L-Arabinofuranosidase (Bifidobacterium adolescentis) for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

Additional Carbohydrate Active enZYme products available.

Certificate of Analysis
Safety Data Sheet
Data Sheet
Megazyme publication
Hydrolysis of wheat flour arabinoxylan, acid-debranched wheat flour arabinoxylan and arabino-xylo-oligosaccharides by β-xylanase, α-L-arabinofuranosidase and β-xylosidase.

McCleary, B. V., McKie, V. A., Draga, A., Rooney, E., Mangan, D. & Larkin, J. (2015). Carbohydrate Research, 407, 79-96.

A range of α-L-arabinofuranosyl-(1-4)-β-D-xylo-oligosaccharides (AXOS) were produced by hydrolysis of wheat flour arabinoxylan (WAX) and acid debranched arabinoxylan (ADWAX), in the presence and absence of an AXH-d3 α-L-arabinofuranosidase, by several GH10 and GH11 β-xylanases. The structures of the oligosaccharides were characterised by GC-MS and NMR and by hydrolysis by a range of α-L-arabinofuranosidases and β-xylosidase. The AXOS were purified and used to characterise the action patterns of the specific α-L-arabinofuranosidases. These enzymes, in combination with either Cellvibrio mixtus or Neocallimastix patriciarum β -xylanase, were used to produce elevated levels of specific AXOS on hydrolysis of WAX, such as 32-α-L-Araf-(1-4)-β-D-xylobiose (A3X), 23-α-L-Araf-(1-4)-β-D-xylotriose (A2XX), 33-α-L-Araf-(1-4)-β-D-xylotriose (A3XX), 22-α-L-Araf-(1-4)-β-D-xylotriose (XA2X), 32-α-L-Araf (1-4)-β-D-xylotriose (XA3X), 23-α-L-Araf-(1-4)-β-D-xylotetraose (XA2XX), 33-α-L-Araf-(1-4)-β-D-xylotetraose (XA3XX), 23 ,33-di-α-L-Araf-(1-4)-β-D-xylotriose (A2+3XX), 23,33-di-α-L-Araf-(1-4)-β-D-xylotetraose (XA2+3XX), 24,34-di-α-L-Araf-(1-4)-β-D-xylopentaose (XA2+3XXX) and 33,34-di-α-L-Araf-(1-4)-β-D-xylopentaose (XA3A3XX), many of which have not previously been produced in sufficient quantities to allow their use as substrates in further enzymic studies. For A2,3XX, yields of approximately 16% of the starting material (wheat arabinoxylan) have been achieved. Mixtures of the α-L-arabinofuranosidases, with specific action on AXOS, have been combined with β-xylosidase and β-xylanase to obtain an optimal mixture for hydrolysis of arabinoxylan to L-arabinose and D-xylose.

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Megazyme publication
Cloning and characterization of arabinoxylan arabinofuranohydrolase-D3 (AXHd3) from Bifidobacterium adolescentis DSM20083.

Van den Broek, L. A. M., Lloyd, R. M., Beldman, G., Verdoes, J. C., McCleary, B. V. & Voragen, A. G. J. (2005). Applied Microbiology and Biotechnology, 67(5), 641-647.

Arabinoxylan arabinofuranohydrolase-D3 (AXHd3) from Bifidobacterium adolescentis releases only C3-linked arabinose residues from double-substituted xylose residues. A genomic library of B. adolescentis DSM20083 was screened for the presence of the axhD3 gene. Two plasmids were identified containing part of the axhD3 gene. The nucleotide sequences were combined and three open reading frames (ORFs) were found. The first ORF showed high homology with xylanases belonging to family 8 of the glycoside hydrolases and this gene was designated xylA. The second ORF was the axhD3 gene belonging to glycoside hydrolase family 43. The third (partial) ORF coded for a putative carboxylesterase. The axhD3 gene was cloned and expressed in Escherichia coli. Several substrates were employed in the biochemical characterization of recombinant AXHd3. The enzyme showed the highest activity toward wheat arabinoxylan oligosaccharides. In addition, β-xylanase from Trichoderma sp. was able to degrade soluble wheat arabinoxylan polymer to a higher extent, after pretreatment with recombinant AXHd3. Arabinoxylan oligosaccharides incubated with a combination of recombinant AXHd3 and an α-L-arabinofuranosidase from Aspergillus niger did not result in a higher maximal release of arabinose than incubation with these enzymes separately.

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Determination of chemical structure of pea pectin by using pectinolytic enzymes.

Noguchi, M., Hasegawa, Y., Suzuki, S., Nakazawa, M., Ueda, M. & Sakamoto, T. (2020). Carbohydrate Polymers, 231, 115738.

The chemical structure of pea pectin was delineated using pectin-degrading enzymes and biochemical methods. The molecular weight of the pea pectin preparation was 488,000, with 50 % arabinose content, and neutral sugar side chains attached to approximately 60 % of the rhamnose residues in rhamnogalacturonan-I (RG-I). Arabinan, an RG-I side chain, was highly branched, and the main chain was comprised of α-1,5-L-arabinan. Galactose and galactooligosaccharides were attached to approximately 35 % of the rhamnose residues in RG-I. Long chain β-1,4-galactan was also present. The xylose substitution rate in xylogalacturonan (XGA) was 63 %. The molar ratio of RG-I/homogalacturonan (HG)/XGA in the backbone of the pea pectin was approximately 3:3:4. When considering neutral sugar side chain content (arabinose, galactose, and xylose), the molar ratio of RG-I/HG/XGA regions in the pea pectin was 7:1:2. These data will help understand the properties of pea pectin.

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Kinetics and regioselectivity of three GH62 α-L-arabinofuranosidases from plant pathogenic fungi.

Sarch, C., Suzuki, H., Master, E. R. & Wang, W. (2019). Biochimica et Biophysica Acta (BBA)-General Subjects, 1863(6), 1070-1078.

Backgound: Xylan is the second most abundant plant cell wall polysaccharide after cellulose with α-L-arabinofuranose (L-Araf) as one of the major side substituents. Capacity to degrade xylan is characteristic of many plant pathogens; and corresponding enzymes that debranch arabinoxylan provide tools to tailor xylan functionality or permit its full hydrolysis. Method: Three GH62_2 family α-arabinofuranosidases (Abfs) from plant pathogenic fungi, NhaAbf62A from Nectria haematococca, SreAbf62A from Sporisorium reilianum and GzeAbf62A from Gibberella zeae, were recombinantly produced in Escherichia coli. Their biochemical properties and substrate specificities were characterized in detail. Particularly with 1H NMR, the regioselectivity and debranching preference of the three Abfs were directly compared. Results: The activities of selected Abfs towards arabinoxylan were all optimal at pH 6.5. Their preferred substrates were wheat arabinoxylan, followed by soluble oat spelt xylan. The Abfs displayed selectivity towards either α-(1 → 2) or α-(1 → 3)-L-Araf mono-substituents in arabinoxylan. Specifically, SreAbf62A and GzeAbf62A removed m-α-(1 → 3)-L-Araf and m-α-(1 → 2)-L-Araf substituents with a similar rates, whereas NhaAbf62A released m-α-(1→ 3)-L-Araf 1.9 times faster than m-α-(1 → 2)-L-Araf. Major conclusions: Building upon the known selectivity of GH62 family α-arabinofuranosidases towards L-Araf mono-substituents in xylans, the current study uncovers enzyme-dependent preferences towards m-α-(1 → 3)-L-Araf and m-α-(1 → 2)-L-Araf substitutions. Comparative sequence-structure analyses of Abfs identified an arginine residue in the xylose binding +2R subsite that was correlated to the observed enzyme-dependent L-Araf debranching preferences. General significance: This study expands the limited pool of characterized GH62 Abfs particularly those from plant pathogenic fungi, and provides biochemical details and methodology to evaluate regioselectivity within this glycoside hydrolase family.

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Constructing arabinofuranosidases for dual arabinoxylan debranching activity.

Wang, W., Andric, N., Sarch, C., Silva, B. T., Tenkanen, M. & Master, E. R. (2017). Biotechnology and Bioengineering, In Press.

Enzymatic conversion of arabinoxylan requires α-L-arabinofuranosidases able to remove α--L-arabinofuranosyl residues (α-L-Araf) from both mono- and double-substituted D-xylopyranosyl residues (Xylp) in xylan (i.e., AXH-m and AXH-d activity). Herein, SthAbf62A (a family GH62 α-L-arabinofuranosidase with AXH-m activity) and BadAbf43A (a family GH43 α-L-arabinofuranosidase with AXH-d3 activity), were fused to create SthAbf62A-BadAbf43A and BadAbf43A-SthAbf62A. Both fusion enzymes displayed dual AXH-m,d and synergistic activity towards native, highly branched wheat arabinoxylan (WAX). When using a customized arabinoxylan substrate comprising mainly α-(1→3)-L-Araf and α-(1→2)-L-Araf substituents attached to disubstituted Xylp (d-2,3-WAX), the specific activity of the fusion enzymes was twice that of enzymes added as separate proteins. Moreover, the SthAbf62A-BadAbf43A fusion removed 83% of all α--L-Araf from WAX after a 20 h treatment. 1H NMR analyses further revealed differences in SthAbf62A-BadAbf43 rate of removal of specific α-L-Araf substituents from WAX, where 9.4 times higher activity was observed towards d-α-(1→3)-L-Araf compared to m-α-(1→3)-L-Araf positions.

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Production of structurally diverse wheat arabinoxylan hydrolyzates using combinations of xylanase and arabinofuranosidase.

Mendis, M. & Simsek, S. (2015). Carbohydrate Polymers, 132, 452-459.

Structurally different wheat arabinoxylan hydrolyzates (AXH) were generated using different combinations of Cellvibrio japonicas xylanase (CJX), Aspergillus niger xylanase (ANX), Bifidobacterium adolescentis arabinofuranosidase (BAF) and Clostridium thermocellum arabinofuranosidase (CAF). Between the two xylanases, ANX might be an enzyme of choice for the production of AXH with simple structural details while CJX might be selected for the production of AXH with more complex structural features. Addition of BAF followed by CAF is more effective in generating AXH with higher amount of unsubstituted xylose. CJX series resulted in lower molecular weights compared to ANX series. The information derived about the capabilities of the two xylanases and two arabinofuranosidase could provide important information in decision making regarding enzymes to be used to generate AXH with specific structural details. Such hydrolyzates could be useful as substrate for future research exploring the effect of fine structural details in AXH on their biological and physical properties.

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Specific enzymatic tailoring of wheat arabinoxylan reveals the role of substitution on xylan film properties.

Heikkinen, S. L., Mikkonen, K. S., Pirkkalainen, K., Serimaa, R., Joly, C. & Tenkanen, M. (2013). Carbohydrate Polymers, 92(1), 733-740.

To increase understanding of the applicability of agro biomass by-products as biodegradable film formers, the effect of wheat arabinoxylan (WAX) fine structure on film properties was studied by applying specific enzyme modifications. WAX was selectively modified to mimic the natural variations of different arabinoxylans, particularly the degree of mono and disubstitution of α-L-arabinofuranosyl (Araf) units in β-D-xylopyranosyl (Xylp) backbone residues. The resulting modified WAX samples had similar arabinose-to-xylose (Ara/Xyl) ratios, but they differed in the number of unsubstituted Xylp units. The substitution of WAX was found to affect, in particular, tensile strength, crystallinity, and oxygen permeability properties of the films, as statistically significant decreases in tensile strength and oxygen permeability took place after WAX de-branching. An increase in the number of unsubstituted Xylp units decreased the temperature of relaxation of small-scale molecular motions of WAX (β-relaxation) and increased the degree of crystallinity of the films.

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Enzyme kinetics and identification of the rate-limiting step of enzymatic arabinoxylan degradation.

Rasmussen, L. E., Xu, C., Sørensen, J. F., Nielsen, M. K. & Meyer, A. S. (2012). Biochemical Engineering Journal, 69, 8-16.

This study investigated the kinetics of multi-enzymatic degradation of soluble wheat arabinoxylan by monitoring the release of xylose and arabinose during designed treatments with mono-component enzymes at different substrate concentrations. The results of different combinations of α-L-arabinofuranosidases (EC, one derived from Aspergillus niger (AFAn) and one from Bifidobacterium adolescentis (AFBa), respectively, a β-xylosidase (EC from Trichoderma reesei, and an engineered D11F/R122D variant of Bacillus subtilis XynA endo-1,4-β-xylanase (EC were examined. The two selected α-L-arabinofuranosidases catalyze liberation of arabinose residues linked 1 → 3 to singly (AFAn) or doubly (AFBa) substituted xyloses in arabinoxylan, respectively. When added to arabinoxylan at equimolar levels, the AFBa enzyme catalyzed the release of more arabinose, i.e. had a higher rate constant than AFAn, but with respect to the xylose release, AFAn– as expected – exhibited a better synergistic effect than AFBa with β-xylosidase. This synergistic effect with AFAn was estimated to increase the number of β-xylosidase catalyzed cuts from ~3 (with β-xylosidase alone) to ~7 in each arabinoxylan substrate molecule. However, the synergistic effects between β-xylosidase and the α-L-arabinofuranosidases on the xylose release were low as compared to the effect of xylanase addition with β-xylosidase, which increased the xylose release by ~25 times in 30 min, to a yield equivalent to ~104 β-xylosidase catalyzed cuts in each arabinoxylan substrate molecule. At equimolar addition levels of the four enzymes, the xylanase activity was thus rate-limiting for the β-xylosidase catalyzed depolymerization to release xylose from arabinoxylan. The work provides clues to design efficient enzymatic degradation of arabinoxylan into fermentable monosaccharides.

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Molecular characterization and solution properties of enzymatically tailored arabinoxylans.

Pitkänen, L., Tuomainen, P., Virkki, L. & Tenkanen, M. (2011). International Journal of Biological Macromolecules, 49(5), 963-969.

Two α-L-arabinofuranosidases with different substrate specificities were used to modify the arabinose-to-xylose ratio of cereal arabinoxylans: one enzyme (AXH-m) removed the L-arabinofuranosyl substituents from the monosubstituted xylopyranosyl residues and the other (AXH-d3) the (1 → 3)-linked L-arabinofuranosyl units from the disubstituted xylopyranosyl residue. In this study, we noticed that not only the arabinose-to-xylose ratio but also the position of the arabinofuranosyl substituents affects the water-solubility of arabinoxylans. The AXH-d3 treatment had no significant effect on the solution conformation of arabinoxylans, but the density of the arabinoxylan molecules decreased in DMSO solution after AXH-m modification. The possible heterogeneity of arabinoxylans complicated the interpretation of data describing the macromolecular properties of the enzymatically modified samples.

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Characterization of indigestible carbohydrates in various fractions from wheat processing.

Haskå, L., Nyman, M. & Andersson, R. (2010). Cereal Chemistry, 87(2), 125-130.

Fractions rich in indigestible carbohydrates, such as fructan and arabinoxylan, are obtained as by-products when ethanol, starch, and gluten are produced from wheat flour. Today, these fractions are used as animal feed. However, these components may have positive physiological effects in humans. In this study, the content of indigestible carbohydrates in distillers' grains and process streams from the wet fractionation of wheat flour was determined. The fractions were further characterized by ethanol extractability analysis, anion-exchange chromatography, NMR, and size-exclusion chromatography. One fraction from wet fractionation contained (g/100 g, db) 6.0 ± 1.0 fructan and 10.3 ± 1.1 dietary fiber (66 ± 4% arabinoxylan), while distillers' grains contained 20.7 g/100 g (db) dietary fiber (30% arabinoxylan). In addition to indigestible carbohydrates from wheat, distillers' grains contained β-(1→3) and β-(1→6) glucans and mannoproteins from the yeast and low molecular weight carbohydrates mainly composed of arabinose. The use of endoxylanase in wet fractionation decreased the molecular weight of the arabinoxylans and increased the arabinose to xylose ratio but had no effect on the fructans. In conclusion, waste streams from industrial wheat processing were enriched in fructan, arabinoxylan, and other indigestible carbohydrates. However, the physiological effects of these fractions require further investigation.

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Safety Data Sheet
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