Bimodal substrate binding in the active site of the glycosidase BcX.

Bacillus circulans xylanase (BcX) from the glycoside hydrolase family 11 degrades xylan through a retaining, double-displacement mechanism. The enzyme is thought to hydrolyze glycosidic bonds in a processive manner and has a large, active site cleft, with six subsites allowing the binding of six xylose units. Such an active site architecture suggests that oligomeric xylose substrates can bind in multiple ways. In the crystal structure of the catalytically inactive variant BcX E78Q, the substrate xylotriose is observed in the active site, as well as bound to the known secondary binding site and a third site on the protein surface. Nuclear magnetic resonance (NMR) titrations with xylose oligomers of different lengths yield nonlinear chemical shift trajectories for active site nuclei resonances, indicative of multiple binding orientations for these substrates for which binding and dissociation are in fast exchange on the NMR timescale, exchanging on the micro- to millisecond timescale. Active site binding can be modeled with a 2 : 1 model with dissociation constants in the low and high millimolar range. Extensive mutagenesis of active site residues indicates that tight binding occurs in the glycon binding site and is stabilized by Trp9 and the thumb region. Mutations F125A and W71A lead to large structural rearrangements. Binding at the glycon site is sensed throughout the active site, whereas the weak binding mostly affects the aglycon site. The interactions with the two active site locations are largely independent of each other and of binding at the secondary binding site.

Bio-bleaching of ankara pulp with xylanase-producing bacterial consortium for sustainable handmade paper production.

The paper industry faces two critical challenges: the scarcity of raw materials and the environmental impact of chemical waste pollution. Addressing the first challenge involves harnessing alternative, sustainable raw materials, while the second challenge can be mitigated through the adoption of bio-bleaching processes, which significantly reduce chemical consumption while enhancing paper brightness and quality. This study proposes a solution to both challenges by using non-woody Calotropis procera (Ankara) and a xylanase-producing microbial consortium for sustainable handmade paper production, a combination not extensively explored in prior research. To evaluate this approach, the process was divided into three stages. In stage I, Ankara fibre was pulped through open hot digestion. In stage II, the pulp was subjected to bio-bleaching in two experimental setups: Set I (without sucrose) and Set II (with sucrose) for 5 days. In stage III, chemical bleaching was used to improve the final brightness of the treated pulps. A novel comparison was made between the bio-bleaching efficiency of an individual isolate g5 (BI) and a bacterial consortium (BC). This research highlighted that bio-bleaching with the consortium effectively removed lignin (140±60 mg/l) and colour (1830±50 PCU), especially in the presence of sucrose, compared to using a single xylanase isolate. Pulp residue/filtrate collected at each stage was estimated based on parameters such as colour and lignin content. After stage III (chemical bleaching), the release of colour and lignin in pulp filtrate was higher in BI compared to BC, indicating the consortium's effectiveness during bio-bleaching, which leaves fewer degradable lignin structures for the chemical bleaching stage. Papers crafted from consortium-treated pulp also exhibited higher brightness than those treated with the isolate. This study reveals the synergistic effect of microbial consortia, leading to more efficient lignin degradation and enhanced bio-bleaching capabilities, supporting the development of greener industrial processes. Ultimately, this study demonstrates a unique and eco-friendly approach to papermaking, combining C. procera and enzymatic bio-bleaching to reduce dependency on hazardous chemicals and support sustainable industry practices.

Production and characterization of acidophilic xylanase from wood degrading white rot fungus by solid-state fermentation of wheat straw.

Xylanases (EC 3.2.1.8) catalyze the breakdown of xylan, which is the second most abundant polysaccharide in plant cell walls. Biological catalysts have gained greater global attention than chemical catalysts in different industrial processes because they are highly selective, easy to control and have a negligible environmental impact. The aim of this study was to investigate the xylanolytic potential of white-rot fungi, optimize their physicochemical conditions and characterize the resulting xylanase. Sixty-eight white-rot fungus (WRF) isolates were screened for their xylanolytic potential and growth conditions for maximal xylanase production using cheap agricultural residue (wheat straw) as the sole carbon source. Five WRF isolates with high xylanase yields (73.63 ± 0.0283-63.6 ± 0.01247 U/ml) were selected by qualitative and quantitative screening methods. The optimum xylanase production occurred at pH 5.0 and 28 °C. Solid-state fermentation (SSF) yielded a high amount of xylanase. The highest xylanase activity (80.9-61.274 U/mL) was recorded in the pH range of 5.0-6.5 and at 50 °C. The metal ions Mg2+, Ca2+ and Mn2+ enhanced the activity of xylanase (127.28-110.06 %), while Cu2+, Fe2+ and K+ inhibited the activity with 43.4-17 % losses. The km and Vmax were 0.32-0.545 mg/mL and 86.95-113.63 µmol/min/mg, respectively. This finding indicates that wheat straw can be used for large-scale xylanase production under SSF conditions. The pH and temperature profiles and stabilities indicate that the xylanase produced in the present study can be applied in food and animal feed industries.

Improving the catalytic activity and thermostability of Aspergillus niger xylanase through computational design.

Xylanase plays the most important role in catalyzing xylan to xylose moieties. GH11 xylanases have been widely used in many fields, but most GH11 xylanases are mesophilic enzymes. To improve the catalytic activity and thermostability of Aspergillus niger xylanase (Xyn-WT), we predicted potential key mutation sites of Xyn-WT through multiple computer-aided enzyme engineering strategies. We introduce a simple and economical Ni affinity chromatography purification method to obtain high-purity xylanase and its mutants. Ten mutants (Xyn-A, Xyn-B, Xyn-C, E45T, Q93R, E45T/Q93R, A161P, Xyn-D, Xyn-E, Xyn-F) were identified. Among the ten mutants, four (Xyn-A, Xyn-C, A161P, Xyn-F) presented improved thermal stability and activity, with Xyn-F(A161P/E45T/Q93R) being the most thermally stable and active. Compared with Xyn-WT, after heat treatment at 55 °C and 60 °C for 10 min, the remaining enzyme activity of Xyn-F was 12 and 6 times greater than that of Xyn-WT, respectively, and Xyn-F was approximately 1.5 times greater than Xyn-WT when not heat treated. The pH adaptation of Xyn-F was also significantly enhanced. In summary, an improved catalytic activity and thermostability of the design variant Xyn-F has been reported.

Upgraded cellulose and xylan digestions for synergistic enhancements of biomass enzymatic saccharification and bioethanol conversion using engineered Trichoderma reesei strains overproducing mushroom LeGH7 enzyme.

Crop straws provide enormous lignocellulose resources transformable for sustainable biofuels and valuable bioproducts. However, lignocellulose recalcitrance basically restricts essential biomass enzymatic saccharification at large scale. In this study, the mushroom-derived cellobiohydrolase (LeGH7) was introduced into Trichoderma reesei (Rut-C30) to generate two desirable strains, namely GH7-5 and GH7-6. Compared to the Rut-C30 strain, both engineered strains exhibited significantly enhanced enzymatic activities, with ß-glucosidases, endocellulases, cellobiohydrolases, and xylanase activities increasing by 113 %, 140 %, 241 %, and 196 %, respectively. By performing steam explosion and mild alkali pretreatments with mature straws of five bioenergy crops, diverse lignocellulose substrates were effectively digested by the crude enzymes secreted from the engineered strains, leading to the high-yield hexoses released for bioethanol production. Notably, the LeGH7 enzyme purified from engineered strain enabled to act as multiple cellulases and xylanase at higher activities, interpreting how synergistic enhancement of enzymatic saccharification was achieved for distinct lignocellulose substrates in major bioenergy crops. Therefore, this study has identified a novel enzyme that is active for simultaneous hydrolyses of cellulose and xylan, providing an applicable strategy for high biomass enzymatic saccharification and bioethanol conversion in bioenergy crops.

Enzymatic production of xylooligosaccharide from lignocellulosic and marine biomass: A review of current progress, challenges, and its applications in food sectors.

Over the last decade, xylooligosaccharides (XOS) have attracted great attentions because of their unique chemical properties and excellent prebiotic effects. Among the current strategies for XOS production, enzymatic hydrolysis is preferred due to its green and safe process, simplicity in equipment, and high control of the degrees of polymerization. This paper comprehensively summarizes various lignocellulosic biomass and marine biomass employed in enzymatic production of XOS. The importance and advantages of enzyme immobilization in XOS production are also discussed. Many novel immobilization techniques for xylanase are presented. In addition, bioinformatics techniques for the mining and designing of new xylanase are also described. Moreover, XOS has exhibited great potential applications in the food industry as diverse roles, such as a sugar replacer, a fat replacer, and cryoprotectant. This review systematically summarizes the current research progress on the applications of XOS in food sectors, including beverages, bakery products, dairy products, meat products, aquatic products, food packaging film, wall materials, and others. It is anticipated that this paper will act as a reference for the further development and application of XOS in food sectors and other fields.

Unleashing the Power of Evolution in Xylanase Engineering: Investigating the Role of Distal Mutation Regulation.

The drive to enhance enzyme performance in industrial applications frequently clashes with the practical limitations of exhaustive experimental screening, underscoring the urgency for more refined and strategic methodologies in enzyme engineering. In this study, xylanase Xyl-1 was used as the model, coupling evolutionary insights with energy functions to obtain theoretical potential mutants, which were subsequently validated experimentally. We observed that mutations in the nonloop region primarily aimed at enhancing stability and also encountered selective pressure for activity. Notably, mutations in this region simultaneously boosted the Xyl-1 stability and activity, achieving a 65% success rate. Using a greedy strategy, mutant M4 was developed, achieving a 12 °C higher melting temperature and doubled activity. By integration of spectroscopy, crystallography, and quantum mechanics/molecular mechanics molecular dynamics, the mechanism behind the enhanced thermal stability of M4 was elucidated. It was determined that the activity differences between M4 and the wild type were primarily driven by dynamic factors influenced by distal mutations. In conclusion, the study emphasizes the pivotal role of evolution-based approaches in augmenting the stability and activity of the enzymes. It sheds light on the unique adaptive mechanisms employed by various structural regions of proteins and expands our understanding of the intricate relationship between distant mutations and enzyme dynamics.

Determination of prebiotic properties of rice bran extract.

This research investigated and compared the prebiotic properties of a rice bran extract obtained through commercial xylanase extraction in comparison with water extraction. Prebiotic properties were evaluated by probiotic growth stimulation (Lacticaseibacillus casei and Lactiplantibacillus plantarum) and gastrointestinal pathogen inhibition (Bacillus cereus and Escherichia coli). The rice bran extract obtained with xylanase (RB1) displayed significantly higher total polysaccharide and total reducing sugar contents than those obtained with water (RB2; p<0.05). After extraction for 30 min, RB1 exhibited the highest total polysaccharide and total reducing sugar contents. HPLC (high performance liquid chromatography) analysis revealed that RB1 primarily contained xylose, while RB2 contained less glucose and lacked other sugar derivatives. RB1 proved effective in stimulating the growth of L. casei and L. plantarum, surpassing even inulin (a commercial prebiotic). Furthermore, it demonstrated a high potential for inhibiting the growth of pathogenic B. cereus and E. coli, comparable to inulin. In contrast, RB2 exhibited lower inhibitory capacity against B. cereus and E. coli.

Enzyme Activity of Culturable Fungi and Bacteria Isolated from Traditional Agarwood Fermentation Basin Indicate Temporally Significant Lignocellulosic and Lipid Substrate Modulations.

Agarwood oil is one of the costliest essential oils used in perfumery, medicine and aroma. Production of the oil traditionally involves a soaking/fermentation step. Studies have indicated a definite role of the diverse microorganisms growing during the open soaking step, and in the emergent aroma of the essential oil. However, the temporal nature of fermentation and a key functional aspect i.e., the enzymatic properties of the microbes from the fermentation basin have not been studied yet. A total of 20 bacteria and 14 fungi isolated from fermentation basins located in Assam, India, at different soaking periods classified as early (0-20 days), medium (20-40 days) and late (40-60 days) clearly pointed towards an early fungal domination followed by succession of bacteria. The physico-chemical transformations of the wood are controlled by enzymatic properties (cellulase, xylanase, amylase and lipase) of the isolates. The results indicated a strong lignocellulosic substrate modulation potential in the four isolates, viz- Purpureocillium lilacinum (0.354 mg/mL), Mucor circinelloides (0.331 mg/mL), Penicillium citrinum (0.324 mg/mL) and Bacillus megaterium (0.152 mg/mL). The highest culturable abundance (CFU/mL) was found in M. circinelloides (2 × 109) among fungi and B. megaterium (4.5 × 109) among bacteria. The highest cellulase activity was shown by P. lilacinum (0.354 mg/mL) while xylanase and lipase by M. circinelloides (0.873 and 0.128 mg/mL). An interesting revelation was that a substantial proportion of the isolates (70% bacteria and 78% fungi) were positive for lipase activity. This is the first report on the "culturable microbiome" of the agarwood fermentation basin from a temporal and functional bioactivity perspective. Supplementary Information: The online version contains supplementary material available at 10.1007/s12088-024-01257-y.

Evolution of glucuronoxylan side chain variability in vascular plants and the compensatory adaptations of cell wall-degrading hydrolases.

Polysaccharide structural complexity not only influences cell wall strength and extensibility but also hinders pathogenic and biotechnological attempts to saccharify the wall. In certain species and tissues, glucuronic acid side groups on xylan exhibit arabinopyranose or galactose decorations whose genetic and evolutionary basis is completely unknown, impeding efforts to understand their function and engineer wall digestibility. Genetics and polysaccharide profiling were used to identify the responsible loci in Arabidopsis and Eucalyptus from proposed candidates, while phylogenies uncovered a shared evolutionary origin. GH30-family endo-glucuronoxylanase activities were analysed by electrophoresis, and their differing specificities were rationalised by phylogeny and structural analysis. The newly identified xylan arabinopyranosyltransferases comprise an overlooked subfamily in the GT47-A family of Golgi glycosyltransferases, previously assumed to comprise mainly xyloglucan galactosyltransferases, highlighting an unanticipated adaptation of both donor and acceptor specificities. Further neofunctionalisation has produced a Myrtaceae-specific xylan galactosyltransferase. Simultaneously, GH30 endo-glucuronoxylanases have convergently adapted to overcome these decorations, suggesting a role for these structures in defence. The differential expression of glucuronoxylan-modifying genes across Eucalyptus tissues, however, hints at further functions. Our results demonstrate the rapid adaptability of biosynthetic and degradative carbohydrate-active enzyme activities, providing insight into plant-pathogen interactions and facilitating plant cell wall biotechnological utilisation.

Co-administration of xylo-oligosaccharides produced by immobilized Aspergillus terreus xylanase with carbimazole to mitigate its adverse effects on the adrenal gland.

Carbimazole has disadvantages on different body organs, especially the thyroid gland and, rarely, the adrenal glands. Most studies have not suggested any solution or medication for ameliorating the noxious effects of drugs on the glands. Our study focused on the production of xylooligosaccharide (XOS), which, when coadministered with carbimazole, relieves the toxic effects of the drug on the adrenal glands. In addition to accelerating the regeneration of adrenal gland cells, XOS significantly decreases the oxidative stress caused by obesity. This XOS produced by Aspergillus terreus xylanase was covalently immobilized using microbial Scleroglucan gel beads, which improved the immobilization yield, efficiency, and operational stability. Over a wide pH range (6-7.5), the covalent immobilization of xylanase on scleroglucan increased xylanase activity compared to that of its free form. Additionally, the reaction temperature was increased to 65 °C. However, the immobilized enzyme demonstrated superior thermal stability, sustaining 80.22% of its original activity at 60 °C for 120 min. Additionally, the full activity of the immobilized enzyme was sustained after 12 consecutive cycles, and the activity reached 78.33% after 18 cycles. After 41 days of storage at 4 °C, the immobilized enzyme was still active at approximately 98%. The immobilized enzyme has the capability to produce xylo-oligosaccharides (XOSs). Subsequently, these XOSs can be coadministered alongside carbimazole to mitigate the adverse effects of the drug on the adrenal glands. In addition to accelerating the regeneration of adrenal gland cells, XOS significantly decreases the oxidative stress caused by obesity.

Temperature-dependent structural changes in xylanase II from Trichoderma longibrachiatum.

Endo-ß-1,4-xylanases degrade heteroxylans that constitute the lignocellulosic plant cell wall. This enzyme is widely used in the food, paper, textile, and biorefinery industries. Temperature affects the optimum activity of xylanase and is an important factor in its application. Various structural analyses of xylanase have been performed, but its structural influence by temperature is not fully elucidated. To better understand the structural influence of xylanase due to temperature, the crystal structure of xylanase II from Trichoderma longibrachiatum (TloXynII) at room and cryogenic temperatures was determined at 2.1 and 1.9 Å resolution, respectively. The room-temperature structure of TloXynII (TloXynIIRT) showed a B-factor value 2.09 times higher than that of the cryogenic-temperature structure of TloXynII (TloXynIICryo). Subtle movement of the catalytic and substrate binding residues was observed between TloXynIIRT and TloXynIICryo. In TloXynIIRT, the thumb domain exhibited high flexibility, whereas in TloXynIICryo, the finger domain exhibited high flexibility. The substrate binding cleft of TloXynIIRT was narrower than that of TloXynIICryo, indicating a distinct finger domain conformation. Numerous water molecule networks were observed in the substrate binding cleft of TloXynIICryo, whereas only a few water molecules were observed in TloXynIIRT. These structural analyses expand our understanding of the temperature-dependent conformational changes in xylanase.

Identification of a novel glycoside hydrolase family 8 xylanase from Deinococcus geothermalis and its application at low temperatures.

Xylanase is the most important hydrolase in the xylan hydrolase system, the main function of which is ß-1,4-endo-xylanase, which randomly cleaves xylans to xylo-oligosaccharides and xylose. Xylanase has wide ranging of applications, but there remains little research on the cold-adapted enzymes required in some low-temperature industries. Glycoside hydrolase family 8 (GH8) xylanases have been reported to have cold-adapted enzyme activity. In this study, the xylanase gene dgeoxyn was excavated from Deinococcus geothermalis through sequence alignment. The recombinant xylanase DgeoXyn encodes 403 amino acids with a theoretical molecular weight of 45.39 kDa. Structural analysis showed that DgeoXyn has a (α/α)6-barrel fold structure typical of GH8 xylanase. At the same time, it has strict substrate specificity, is only active against xylan, and its hydrolysis products include xylobiose, xylotrinose, xytetranose, xylenanose, and a small amount of xylose. DgeoXyn is most active at 70 â„ƒ and pH 6.0. It is very stable at 10, 20, and 30 â„ƒ, retaining more than 80% of its maximum enzyme activity. The enzyme activity of DgeoXyn increased by 10% after the addition of Mn2+ and decreased by 80% after the addition of Cu2+. The Km and Vmax of dgeox were 42 mg/ml and 20,000 U/mg, respectively, at a temperature of 70 â„ƒ and pH of 6.0 using 10 mg/ml beechwood xylan as the substrate. This research on DgeoXyn will provide a theoretical basis for the development and application of low-temperature xylanase.

Advances in the understanding of the production, modification and applications of xylanases in the food industry.

Xylanases have broad applications in the food industry to decompose the complex carbohydrate xylan. This is applicable to enhance juice clarity, improve dough softness, or reduce beer turbidity. It can also be used to produce prebiotics and increase the nutritional value in foodstuff. However, the low yield and poor stability of most natural xylanases hinders their further applications. Therefore, it is imperative to explore higher-quality xylanases to address the potential challenges that appear in the food industry and to comprehensively improve the production, modification, and utilization of xylanases. Xylanases, due to their various sources, exhibit diverse characteristics that affect production and activity. Most fungi are suitable for solid-state fermentation to produce xylanases, but in liquid fermentation, microbial metabolism is more vigorous, resulting in higher yield. Fungi produce higher xylanase activity, but bacterial xylanases perform better than fungal ones under certain extreme conditions (high temperature, extreme pH). Gene and protein engineering technology helps to improve the production efficiency of xylanases and enhances their thermal stability and catalytic properties.

A new halotolerant xylanase from Aspergillus clavatus expressed in Escherichia coli with catalytic efficiency improved by site-directed mutagenesis.

Daily agro-industrial waste, primarily cellulose, lignin, and hemicellulose, poses a significant environmental challenge. Harnessing lignocellulolytic enzymes, particularly endo-1,4-ß-xylanases, for efficient saccharification is a cost-effective strategy, transforming biomass into high-value products. This study focuses on the cloning, expression, site-directed mutagenesis, purification, three-dimensional modeling, and characterization of the recombinant endo-1,4-ß-xylanase (XlnA) from Aspergillus clavatus in Escherichia coli. This work includes evaluation of the stability at varied NaCl concentrations, determining kinetic constants, and presenting the heterologous expression of XlnAΔ36 using pET22b(+). The expression led to purified enzymes with robust stability across diverse pH levels, exceptional thermostability at 50 °C, and 96-100% relative stability after 24 h in 3.0 M NaCl. Three-dimensional modeling reveals a GH11 architecture with catalytic residues Glu 132 and 22. XlnAΔ36 demonstrates outstanding kinetic parameters compared to other endo-1,4-ß-xylanases, indicating its potential for industrial enzymatic cocktails, enhancing saccharification. Moreover, its ability to yield high-value compounds, such as sugars, suggests a promising and ecologically positive alternative for the food and biotechnology industries.

Effect of Air Classification and Enzymatic and Microbial Bioprocessing on Defatted Durum Wheat Germ: Characterization and Use as Bread Ingredient.

Its high dietary fiber and protein contents and nutritional quality make defatted wheat germ (DWG) a valuable cereal by-product, yet its negative impact on food structure limits its use as a food ingredient. In this research, DWG underwent air classification, which identified two fractions with high fiber (HF) and low fiber/high protein (LF) contents, and a bioprocessing protocol, involving treatment with xylanase and fermentation with selected lactic acid bacterial strains. The degree of proteolysis was evaluated through electrophoretic and chromatographic techniques, revealing differences among fractions and bioprocessing options. Fermentation led to a significant increase in free amino acids (up to 6 g/kg), further enhanced by the combination with xylanase. When HF was used as an ingredient in bread making, the fiber content of the resulting bread exceeded 3.6 g/100 g, thus reaching the threshold required to make a "source of fiber" claim according to Regulation EC No.1924/2006. Meanwhile, all breads could be labeled a "source of protein" since up to 13% of the energy was provided by proteins. Overall, bioprocessed ingredients lowered the glycemic index (84 vs. 89) and increased protein digestibility (80 vs. 63%) compared to control breads. Technological and sensory analysis showed that the enzymatic treatment combined with fermentation also conferred a darker and more pleasant color to the bread crust, as well as better crumb porosity and elasticity.

A novel xylanase from a myxobacterium triggers a plant immune response in Nicotiana benthamiana.

Xylanases derived from fungi, including phytopathogenic and nonpathogenic fungi, are commonly known to trigger plant immune responses. However, there is limited research on the ability of bacterial-derived xylanases to trigger plant immunity. Here, a novel xylanase named CcXyn was identified from the myxobacterium Cystobacter sp. 0969, which displays broad-spectrum activity against both phytopathogenic fungi and bacteria. CcXyn belongs to the glycoside hydrolases (GH) 11 family and shares a sequence identity of approximately 32.0%-45.0% with fungal xylanases known to trigger plant immune responses. Treatment of Nicotiana benthamiana with purified CcXyn resulted in the induction of hypersensitive response (HR) and defence responses, such as the production of reactive oxygen species (ROS) and upregulation of defence gene expression, ultimately enhancing the resistance of N. benthamiana to Phytophthora nicotianae. These findings indicated that CcXyn functions as a microbe-associated molecular pattern (MAMP) elicitor for plant immune responses, independent of its enzymatic activity. Similar to fungal xylanases, CcXyn was recognized by the NbRXEGL1 receptor on the cell membrane of N. benthamiana. Downstream signalling was shown to be independent of the BAK1 and SOBIR1 co-receptors, indicating the involvement of other co-receptors in signal transduction following CcXyn recognition in N. benthamiana. Moreover, xylanases from other myxobacteria also demonstrated the capacity to trigger plant immune responses in N. benthamiana, indicating that xylanases in myxobacteria are ubiquitous in triggering plant immune functions. This study expands the understanding of xylanases with plant immune response-inducing properties and provides a theoretical basis for potential applications of myxobacteria in biocontrol strategies against phytopathogens.

Investigation of the nutritional and functional roles of a combinational use of xylanase and ß-glucanase on intestinal health and growth of nursery pigs.

BACKGROUND: Xylanase and ß-glucanase combination (XG) hydrolyzes soluble non-starch polysaccharides that are anti-nutritional compounds. This study aimed to evaluate the effects of increasing levels of XG on intestinal health and growth performance of nursery pigs. METHODS: Forty pigs (6.5 ± 0.4 kg) were assigned to 5 dietary treatments and fed for 35 d in 3 phases (11, 9, and 15 d, respectively). Basal diets mainly included corn, soybean meal, and corn distiller's dried grains with solubles, contained phytase (750 FTU/kg), and were supplemented with 5 levels of XG at (1) 0, (2) 280 TXU/kg xylanase and 125 TGU/kg ß-glucanase, (3) 560 and 250, (4) 840 and 375, or (5) 1,120 and 500, respectively. Growth performance was measured. On d 35, all pigs were euthanized and jejunal mucosa, jejunal digesta, jejunal tissues, and ileal digesta were collected to determine the effects of increasing XG levels and XG intake on intestinal health. RESULTS: Increasing XG intake tended to quadratically decrease (P = 0.059) viscosity of jejunal digesta (min: 1.74 mPa·s at 751/335 (TXU/TGU)/kg). Increasing levels of XG quadratically decreased (P < 0.05) Prevotellaceae (min: 0.6% at 630/281 (TXU/TGU)/kg) in the jejunal mucosa. Increasing XG intake quadratically increased (P < 0.05) Lactobacillaceae (max: 40.3% at 608/271 (TXU/TGU)/kg) in the jejunal mucosa. Increasing XG intake quadratically decreased (P < 0.05) Helicobacteraceae (min: 1.6% at 560/250 (TXU/TGU)/kg) in the jejunal mucosa. Increasing levels of XG tended to linearly decrease (P = 0.073) jejunal IgG and tended to quadratically increase (P = 0.085) jejunal villus height to crypt depth ratio (max: 2.62 at 560/250 (TXU/TGU)/kg). Increasing XG intake tended to linearly increase the apparent ileal digestibility of dry matter (P = 0.087) and ether extract (P = 0.065). Increasing XG intake linearly increased (P < 0.05) average daily gain. CONCLUSIONS: A combinational use of xylanase and ß-glucanase would hydrolyze the non-starch polysaccharides fractions, positively modulating the jejunal mucosa-associated microbiota. Increased intake of these enzyme combination possibly reduced digesta viscosity and humoral immune response in the jejunum resulting in improved intestinal structure, and ileal digestibility of nutrients, and finally improving growth of nursery pigs. The beneficial effects were maximized at a combination of 550 to 800 TXU/kg xylanase and 250 to 360 TGU/kg ß-glucanase.

Xylanase VmXyl2 is involved in the pathogenicity of Valsa mali by regulating xylanase activity and inducing cell necrosis.

Xylanase plays a key role in degrading plant cell wall during pathogenic fungi infection. Here, we identified a xylanase gene, VmXyl2 from the transcriptome of Valsa mali and examined its function. VmXyl2 has highly elevated transcript levels during the infection process of V. mali, with 15.02-fold increase. Deletion mutants of the gene were generated to investigate the necessity of VmXyl2 in the development and pathogenicity of V. mali. The VmXyl2 deletion mutant considerably reduced the virulence of V. mali in apple leaves and in twigs, accompanied by 41.22% decrease in xylanase activity. In addition, we found that VmXyl2 induces plant cell necrosis regardless of its xylanase activity, whereas promoting the infection of V. mali in apple tissues. The cell death-inducing activity of VmXyl2 dependent on BRI1-associated kinase-1 (BAK1) but not Suppressor of BIR1-1 (SOBIR1). Furthermore, VmXyl2 interacts with Mp2 in vivo, a receptor-like kinase with leucine-rich repeat. The results offer valuable insights into the roles of VmXyl2 in the pathogenicity of V. mali during its infection of apple trees.