|Formulation:||In 3.2 M ammonium sulphate|
|Stability:||> 4 years at 4oC|
|Synonyms:||endo-1,4-beta-xylanase; 4-beta-D-xylan xylanohydrolase|
|Concentration:||Supplied at ~ 1,600 U/mL|
|Expression:||Purified from Trichoderma longibrachiatum|
|Specificity:||endo-hydrolysis of (1,4)-β-D-xylosidic linkages in xylans.|
|Specific Activity:||> 100 U/mg (40oC, pH 6.0 on wheat arabinoxylan)|
|Unit Definition:||One Unit of xylanase activity is defined as the amount of enzyme required to release one µmole of xylose reducing-sugar equivalents per minute from wheat arabinoxylan (10 mg/mL) in sodium phosphate buffer (100 mM), pH 6.0 at 40oC.|
|Application examples:||Applications in carbohydrate and biofuels research and in the food and feeds and paper pulping industries.|
High purity endo-1,4-β-Xylanase M3 (Trichoderma longibrachiatum) for use in research, biochemical enzyme assays and in vitro diagnostic analysis.
See our entire product listing of Carbohydrate Active enZYmes.
(Bacillus stearothermophilus T6) E-XYNACJ - endo-1,4-β-Xylanase (Cellvibrio japonicus) E-XYLNP - endo-1,4-β-Xylanase (Neocallimastix patriciarum) E-XYLATM - endo-1,4-β-Xylanase (Thermotoga maritima)
Mangan, D., Cornaggia, C., Liadova, A., McCormack, N., Ivory, R., McKie, V. A., Ormerod, A. & McCleary, D. V. (2017). Carbohydrate Research, 445, 14-22.
endo-1,4-β-Xylanase (EC 184.108.40.206) is employed across a broad range of industries including animal feed, brewing, baking, biofuels, detergents and pulp (paper). Despite its importance, a rapid, reliable, reproducible, automatable assay for this enzyme that is based on the use of a chemically defined substrate has not been described to date. Reported herein is a new enzyme coupled assay procedure, termed the XylX6 assay, that employs a novel substrate, namely 4,6-O-(3-ketobutylidene)-4-nitrophenyl-β-45-O-glucosyl-xylopentaoside. The development of the substrate and associated assay is discussed here and the relationship between the activity values obtained with the XylX6 assay versus traditional reducing sugar assays and its specificity and reproducibility were thoroughly investigated.Hide Abstract
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.Hide Abstract
Multimodular fused acetyl-feruloyl esterases from soil and gut Bacteroidetes improve xylanase depolymerization of recalcitrant biomass.
Kmezik, C., Bonzom, C., Olsson, L., Mazurkewich, S., & Larsbrink, J. (2020). Biotechnology for Biofuels, 13, 1-14.
Background: Plant biomass is an abundant and renewable carbon source that is recalcitrant towards both chemical and biochemical degradation. Xylan is the second most abundant polysaccharide in biomass after cellulose, and it possesses a variety of carbohydrate substitutions and non-carbohydrate decorations which can impede enzymatic degradation by glycoside hydrolases. Carbohydrate esterases are able to cleave the ester-linked decorations and thereby improve the accessibility of the xylan backbone to glycoside hydrolases, thus improving the degradation process. Enzymes comprising multiple catalytic glycoside hydrolase domains on the same polypeptide have previously been shown to exhibit intramolecular synergism during degradation of biomass. Similarly, natively fused carbohydrate esterase domains are encoded by certain bacteria, but whether these enzymes can result in similar synergistic boosts in biomass degradation has not previously been evaluated. Results: Two carbohydrate esterases with similar architectures, each comprising two distinct physically linked catalytic domains from families 1 (CE1) and 6 (CE6), were selected from xylan-targeting polysaccharide utilization loci (PULs) encoded by the Bacteroidetes species Bacteroides ovatus and Flavobacterium johnsoniae. The full-length enzymes as well as the individual catalytic domains showed activity on a range of synthetic model substrates, corn cob biomass, and Japanese beechwood biomass, with predominant acetyl esterase activity for the N-terminal CE6 domains and feruloyl esterase activity for the C-terminal CE1 domains. Moreover, several of the enzyme constructs were able to substantially boost the performance of a commercially available xylanase on corn cob biomass (close to twofold) and Japanese beechwood biomass (up to 20-fold). Interestingly, a significant improvement in xylanase biomass degradation was observed following addition of the full-length multidomain enzyme from B. ovatus versus the addition of its two separated single domains, indicating an intramolecular synergy between the esterase domains. Despite high sequence similarities between the esterase domains from B. ovatus and F. johnsoniae, their addition to the xylanolytic reaction led to different degradation patterns. Conclusion: We demonstrated that multidomain carbohydrate esterases, targeting the non-carbohydrate decorations on different xylan polysaccharides, can considerably facilitate glycoside hydrolase-mediated hydrolysis of xylan and xylan-rich biomass. Moreover, we demonstrated for the first time a synergistic effect between the two fused catalytic domains of a multidomain carbohydrate esterase.Hide Abstract
Scarafoni, A., Consonni, A., Pessina, S., Balzaretti, S., Capraro, J., Galanti, E. & Duranti, M. (2016). Plant Physiology and Biochemistry, 99, 79-85.
Lupin γ-conglutin and soybean BG7S are two legume seed proteins strongly similar to plant endo-β-glucanases inhibitors acting against fungal GH11 and GH12 glycoside hydrolase. However these proteins lack inhibitory activity. Here we describe the conversion of lupin γ-conglutin to an active inhibitor of endo-β-glucanases belonging to GH11 family. A set of γ-conglutin mutants was designed and expressed in Pichia pastoris, along with the wild-type protein. Unexpectedly, this latter was able to inhibit a GH11 enzyme, but not GH12, whereas the mutants were able to modulate the inhibition capacity. In lupin, γ-conglutin is naturally cleaved in two subunits, whereas in P. pastoris it is not. The lack of proteolytic cleavage is one of the reasons at the basis of the inhibitory activity of recombinant γ-conglutin. The results provide new insights about structural features at the basis of the lack of inhibitory activity of wild-type γ-conglutin and its legume homologues.Hide Abstract
Tundo, S., Moscetti, I., Faoro, F., Lafond, M., Giardina, T., Favaron, F., Sella, L. & D'Ovidio, R. (2015). Plant Science, 240, 161-169.
To shed light on the role of Xylanase Inhibitors (XIs) during Fusarium graminearum infection, we first demonstrated that three out of four F. graminearum xylanases, in addition to their xylan degrading activity, have also the capacity to cause host cell death both in cell suspensions and wheat spike tissue. Subsequently, we demonstrated that TAXI-III and XIP-I prevented both the enzyme and host cell death activities of F. graminearum xylanases. In particular, we showed that the enzymatic inhibition by TAXI-III and XIP-I was competitive and only FGSG_11487 escaped inhibition. The finding that TAXI-III and XIP-I prevented cell death activity of heat inactivated xylanases and that XIP-I precluded the cell death activity of FGSG_11487 – even if XIP-I does not inhibit its enzyme activity – suggests that the catalytic and the cell death activities are separated features of these xylanases. Finally, the efficacy of TAXI-III or XIP-I to prevent host cell death caused by xylanases was confirmed in transgenic plants expressing separately these inhibitors, suggesting that the XIs could limit F. graminearum infection via direct inhibition of xylanase activity and/or by preventing host cell death.Hide Abstract
Cuyvers, S., Dornez, E., Moers, K., Pollet, A., Delcour, J. A. & Courtin, C. M. (2011). Enzyme and Microbial Technology, 49(3), 305-311.
In biomass degradation using simultaneous saccharification and fermentation (SSF), there is a need for efficient biomass degrading enzymes that can work at lower temperatures suitable for yeast fermentation. As xylan is an important lignocellulosic biomass constituent, this study aimed at investigating the possible differences in xylan breakdown potential of endoxylanases using eight different endoxylanases at conditions relevant for SSF. Both solubilising and degrading capacities of the endoxylanases were investigated using water-insoluble and water-soluble oat spelt xylan as model substrates for biomass xylan. Results showed that selecting for combinations of endoxylanases that are efficient at solubilising xylan on the one hand and degrading it to large extent on the other hand, coupled to high specific activities, seems the best option for complete xylan breakdown in lignocellulosic biomass conversion using SSF.Hide Abstract
Tokunaga, T. & Esaka, M. (2007). Plant and Cell Physiology, 48(5), 700-714.
Rice microarray analysis showed that a number of stress-related genes are induced by external addition of L-ascorbic acid (AsA). The gene designated as AK07384 which is homologous to class Ш chitinase was found to exhibit the highest induction among these genes. However, its crucial residues within the chitinase active site are substituted with other residues, suggesting that the protein has no chitinase activity. The recombinant protein which is encoded by the AK073843 gene produced in Escherichia coli has xylanase inhibitor activity, indicating that the gene encodes a novel rice XIP-type xylanase inhibitor protein (OsXIP). The expression of OsXIP was enhanced not only by exogenous AsA treatment but also by various stresses such as citrate and sodium chloride treatments, and wounding; however, it was not influenced by increasing endogenous AsA content. External AsA treatment caused a significant increase in electrolyte leakage from rice root. These results suggested that OsXIP was induced by stress which is caused by external AsA treatment. Rice XIP-family genes, OsXIP, riceXIP and RIXI, showed differential organ-specific expression. Also, these genes were differentially induced by stress and stress-related phytohormones. The transcripts of OsXIP and riceXIP were undetectable under normal conditions, and were drastically induced by wounding and methyl jasmonate (MeJA) treatment in the root. RIXI was constitutively expressed in the shoot but not induced by wounding and stress-related phytohormones. Thus, XIP-type xylanase inhibitors were suggested to be specialized in their function and involved in defense mechanisms in rice.Hide Abstract
Fierens, K., Gils, A., Sansen, S., Brijs, K., Courtin, C. M., Declerck, P. J., De Ranter, C. J., Gebruers, K., Rabijns, A., Robben, J., Van Campenhout, S., Volckaert, G. & Delcour, J. A. (2005). FEBS Journal, 272(22), 5872-5882.
Wheat endoxylanase inhibitor TAXI-I inhibits microbial glycoside hydrolase family 11 endoxylanases. Crystallographic data of an Aspergillus niger endoxylanase-TAXI-I complex showed His374 of TAXI-I to be a key residue in endoxylanase inhibition [Sansen S, De Ranter CJ, Gebruers K, Brijs K, Courtin CM, Delcour JA & Rabijns A (2004) J Biol Chem 279, 36022–36028]. Its role in enzyme–inhibitor interaction was further investigated by site-directed mutagenesis of His374 into alanine, glutamine or lysine. Binding kinetics and affinities of the molecular interactions between A. niger, Bacillus subtilis, Trichoderma longibrachiatum endoxylanases and wild-type TAXI-I and TAXI-I His374 mutants were determined by surface plasmon resonance analysis. Enzyme–inhibitor binding was in accordance with a simple 1 : 1 binding model. Association and dissociation rate constants of wild-type TAXI-I towards the endoxylanases were in the range between 1.96 and 36.1 × 104m-1·s-1 and 0.72–3.60 × 10-4·s-1, respectively, resulting in equilibrium dissociation constants in the low nanomolar range. Mutation of TAXI-I His374 to a variable degree reduced the inhibition capacity of the inhibitor mainly due to higher complex dissociation rate constants (three- to 80-fold increase). The association rate constants were affected to a smaller extent (up to eightfold decrease). Substitution of TAXI-I His374 therefore strongly affects the affinity of the inhibitor for the enzymes. In addition, the results show that His374 plays a critical role in the stabilization of the endoxylanase–TAXI-I complex rather than in the docking of inhibitor onto enzyme.Hide Abstract
ESEM Study of the Effects of Hydrolytic Enzymes on Wheat Bran Structure.
Douge, M., Nonus, M., Thomasset, T., Teissier, P. & Barbeau, J. Y. (2004). Microscopy and Analysis, 18(6), 21-24.
Wheat bran is a major dietary fibre source, comprising mainly hemicelluloses of the arabinoxylan type, and cellulose. Environmental scanning electron microscopy was used to associate wheat bran solubilisation by enzymatic treatments (cellulase, cellobiohydrolase, α-glucosidase, xylanase, arabinofuranosidase, α-xylosidase, -amylase, α-amylase, amyloglucosidase, -glucosidase) with tissue structure modifications. When cellulase or xylanase were used alone or in association with other enzymes, separation of outer layers (epicarp and mesocarp) from the endocarp and aleurone layers was observed. Starch granule removal was observed only with a cocktail of enzymes.Hide Abstract