Xylanase Activity Research Articles

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.

Transcriptional activator UvXlnR is required for conidiation and pathogenicity of rice false smut fungus Ustilaginoidea virens.

Transcription factors play critical roles in diverse biological processes in fungi. XlnR, identified as a transcriptional activator that regulates the expression of the extracellular xylanase genes in fungi, has not been extensively studied for its function in fungal development and pathogenicity in rice false smut fungus Ustilaginoidea virens. In this study, we characterized UvXlnR in U. virens and established that the full-length, N- and C-terminal forms of the UvXlnR have the ability to activate transcription. The study further demonstrated that UvXlnR plays crucial roles in various aspects of U. virens biology. Deletion of UvXlnR affected growth, conidiation, and stress response. UvXlnR mutants also exhibited reduced pathogenicity, which could be partially attributed to the reduced expression of xylanolytic genes and extracellular xylanase activity of U. virens during the infection process. Our results indicate that UvXlnR is involved in regulating growth, conidiation, stress response, and pathogenicity.

Isolation, Screening and Characterization of Xylose-Fermenting Yeasts Isolated from Saw Dust

Yeasts have been less frequently reported as xylanase producers compared to bacteria and filamentous fungi. Different cellulosic materials including sawdust are produced on a large scale and these can be used for the production of useful enzymes such as xylanases. Xylanases are a group of enzymes that hydrolyze plant fibers made of xylan hemicellulose. Xylose-fermenting yeasts isolated from soil at a wood processing factory were isolated and qualitatively and quantitatively screened for xylanase production using xylose supplemented medium and congo red as indicator. Xylanase enzyme was produced using different xylose concentrations (0.5%, 1.0%, 1.5% and 2.0%). Pichia chambardii isolate, which was later identified as Wickerhamomyces chambardii by molecular techniques, showed the highest xylanase activity of 199.31U mL-1. Maximum xylanase activity (275.83U mL-1) was achieved at 1.5 %w/v xylose. This study showed that yeasts have a high potential for the production of xylanase enzymes.

Effects of Fungal Solid-State Fermentation on the Profile of Phenolic Compounds and on the Nutritional Properties of Grape Pomace.

Grape pomace (GP) is considered a natural source of bioactive compounds. To improve the extractability of bioactive compounds, in this work, GP was biologically treated for 15 days with the white-rot fungus Trametes versicolor in laboratory jars and a tray bioreactor under solid-state fermentation (SSF) conditions. During SSF, the activity of lignolytic (laccase and manganese peroxidase) and hydrolytic (xylanase, cellulase, β-glucosidase, and invertase) enzymes was measured, with the activities of laccase (2.66 U/gdb in jars and 0.96 U/gdb in the bioreactor) and xylanase (346.04 U/gdb in jars and 200.65 U/gdb in the bioreactor) being the highest. The effect of the complex enzyme system was reflected in the changes in the chemical composition of GP with increasing ash, crude protein, and free fat content: 28%, 10%, and 17% in the laboratory jars, and 29%, 11%, and 7% in the bioreactor, respectively. In addition, the biological treatment improved the extractability of 13 individual phenolic compounds. Therefore, the applied SSF technique represents an effective strategy to improve the profile of phenolic compounds and the nutritional composition of GP, promoting their valorization and opening the door for potential applications in the food industry and other sectors.

An efficient preparation process of sisal fibers via the specialized retting microorganisms: Based on the ideal combination of degumming-related enzymes for the effective removal of non-cellulosic macromolecules

Bioaugmentation retting with the specialized pectinolytic and xylanolytic microorganisms can accelerate the removal of non-cellulosic macromolecules around plant fibers, thus shortening retting time and facilitating fiber quality. Currently, few specialized microorganisms have been explored for the retting of sisal fibers. The present study excavated the retting fungi including Aspergillus micronesiensis HD 3–6, Penicillium citrinum HD 3–12-3, and Cladosporium sp. HD 4–13 from the region-specific soil samples of planting sisal, and investigated their bioaugmentation retting effects on raw sisal leaves. Results showed that combination of the three fungi achieved the most excellent degumming efficiency (13.69 % of residual gum in sisal fibers) and the highest fiber yield (4.47 %). Furthermore, this fungi combination had the ideal enzymatic hydrolysis features with high activities of pectinase, xylanase and mannanase whereas a low activity of cellulase during the whole retting process, thus endowing the prepared sisal fibers with the lowest mass percentage of non-cellulosic macromolecules (9.76 wt%) and the highest cellulose content (89.23 wt%). SEM and FT-IR analysis further verified that the non-cellulosic substances around sisal fibers were efficiently removed. In summary, the consortia of the three fungi achieved ideal degumming-related enzymes for the removal of non-cellulosic macromolecules, thus acquiring the efficient preparation of sisal fibers.

Production of xylanase intended to be used in bread-making: laboratory scale and pilot scale studies

In this study, submerged and solid-state fermentation studies were performed for xylanase production simultaneously to determine ideal carbon and nitrogen sources together with necessary production parameters from the laboratory scale to the pilot scale. The results showed that barley malt sprout is effective as a nitrogen and a good carbon source. Under optimum conditions, xylanase activities 71.74 ± 2.70 U/mL, 56.95 ± 3.6 U/mL, 65.8 ± 8 U/mL and 46.05 ± 0.1 U/mL were achieved in shake flasks, 2, 10 and 30 L bioreactors, respectively. In solid-state fermentation studies, maximum activity in flasks was 759.15 ± 36.16 U/g production media, and maximum activity in pilot-scale tray bioreactor productions was 1312 ± 137.85 U/g dry substrate. Bread-making trials showed that using xylanase at 100 U/100g flour resulted in a significant increase in the loaf volume of wheat flour bread and whole wheat flour bread.

Optimization Production of an Endo-β-1,4-Xylanase from Streptomyces thermocarboxydus Using Wheat Bran as Sole Carbon Source

Xylanases, key enzymes for hydrolyzing xylan, have diverse industrial applications. The bioprocessing of agricultural byproducts to produce xylanase through fermentation approaches is gaining importance due to its significant potential to reduce enzyme production costs. In this work, the productivity of Streptomyces thermocarboxydus TKU045 xylanase was enhanced through liquid fermentation employing wheat bran as the sole carbon source. The maximum xylanase activity (25.314 ± 1.635 U/mL) was obtained using the following optima factors: 2% (w/v) wheat bran, 1.4% (w/v) KNO3, an initial pH of 9.8, an incubation temperature of 37.3 °C, and an incubation time of 2.2 days. Xylanase (Xyn_TKU045) of 43 kDa molecular weight was isolated from the culture supernatant and was biochemically characterized. Analysis through liquid chromatography with tandem mass spectrometry revealed a maximum amino acid identity of 19% with an endo-1,4-β-xylanase produced by Streptomyces lividans. Xyn_TKU045 exhibited optimal activity at pH 6, with remarkable stability within the pH range of 6.0 to 8.0. The enzyme demonstrated maximum efficiency at 60 °C and considerable stability at ≤70 °C. Mg2+, Mn2+, Ba2+, Ca2+, 2-mercaptoethanol, Tween 20, Tween 40, and Triton X-100 positively influenced Xyn_TKU045, while Zn2+, Fe2+, Fe3+, Cu2+, and sodium dodecyl sulfate exhibited adverse impact. The kinetic properties of Xyn_TKU045 were a Km of 0.628 mg/mL, a kcat of 75.075 s−1 and a kcat/Km of 119.617 mL mg−1s−1. Finally, Xyn_TKU045 could effectively catalyze birchwood xylan into xylotriose and xylobiose as the major products.

Carbon fixation mechanism of fungal community bypass strategy in rice straw composting combined with functional microbes: inhibition from cellobiose to glucose pathway

Composting offers an eco-friendly approach, converting organic solid waste into valuable soil enhancer products. Three experimental treatments were designed to explore the microbial mechanisms influencing lignocellulose loss in rice straw composting: CK (the control), B4 (inoculated with Bacillus subtilis), and Z1 (inoculated with Aspergillus fumigatus). The results showed significant increases in carboxymethyl cellulase, cellobiohydrolase, laccase, and xylanase activities during B4 and Z1composting. However, lignin peroxidase and manganese peroxidase were not significantly affected, and β-glucosidase was inhibited. Additionally, microbial inoculation stimulated lignin loss, with B4 and Z1 composting increasing degradation by 14.42% and 16.29%, respectively, compared to the CK. Simultaneously, humic substance content increased as microbial inoculation inhibited β-glucosidase, causing cellobiose to accumulate and divert into a pathway that forms humic substances. The random forest model showed that Symmetrospora and Phaeosphaeria significantly boosted lignocellulose loss in B4 and Z1 composting by promoting fungi functional aggregation, thus accelerating the degradation and transformation process. This study explored the microbial mechanism of lignocellulose loss and provided new insights into lignocellulose conversion and carbon fixation.

Enhancement of biochemical parameters and enzyme activity in solid-state fermented and biofortified maize cobs utilizing yeasts and plant extracts

This study investigated the impact of solid-state fermentation and biofortification of maize cobs using yeast and plant extracts on their biochemical properties and enzyme activity. Maize cobs were fermented with extracts from elephant grass, cassava leaves, or Siam weed, combined with baker's yeast or palm wine yeast. In vitro assays revealed increased l-lysine concentration (up to 8.6 mg/g), soluble protein content (up to 27.4 mg/g), total phenolic content (up to 15.5 μg GAE/g), total flavonoid content (up to 23.4 μg CE/g), and xylanase activity (up to 25.7 Units) compared to controls. Glucose concentration decreased (to 0.8 mg/mL), indicating efficient utilization. Amylase and protease activities varied, with some combinations showing higher enzymatic activity (amylase at 35.4 Units, protease at 39.2 Units). These results underscore the potential of solid-state fermentation and biofortification to enhance the nutritional quality of agro-residues, offering novel insights into sustainable food production and waste utilization strategies.

Temperature-dependent structural changes in xylanase II from Trichoderma longibrachiatum

Endo-β-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.

Valorization of Gelidium corneum by-product through solid-state fermentation

Red seaweed Gelidium is used for agar extraction, generating large quantities of by-products (red seaweed by-product – GBP). Its carbohydrate and protein-rich composition makes it a suitable substrate to produce value-added compounds through solid-state fermentation (SSF). For the first time, GBP was used in SSF with Aspergillus ibericus and Aspergillus niger for carbohydrases production and protein enrichment. SSF was performed with unsupplemented GBP, supplemented with Mandel salt solution, and mixed at equal dry mass proportion with plant feedstuffs and with the seaweed Ulva rigida. A. niger CECT 2915 was the best strain under SSF obtaining the maximum xylanase activity (498 ± 49 U g-1), in GBP mixed with sunflower cake and the maximum of cellulase activity (382 ± 37 U g-1) in GBP mixed with rapeseed cake. In these mixtures, protein content increased 1.2-fold by SSF, as well as in GBP mixed with rice bran (1.3-fold). Scale-up of SSF using GBP mixed with sunflower cake from flasks to tray-type bioreactors demonstrated SSF reproducibility, achieving similar xylanase and cellulase activities. In the stirred-drum bioreactor, aeration and low agitation favored both enzymes production. Bioprocessing of GBP with plant feedstuffs using SSF was a cost-effective strategy to produce high-value enzymes, valorizing this seaweed by-product.

Unlocking the potential of Algerian lignocellulosic biomass: exploring indigenous microbial diversity for enhanced enzyme and sugar production.

Nowadays, natural resources like lignocellulosic biomass are gaining more and more attention. This study was conducted to analyse chemical composition of dried and ground samples (500μm) of various Algerian bioresources including alfa stems (AS), dry palms (DP), olive pomace (OP), pinecones (PC), and tomato waste (TW). AS exhibited the lowest lignin content (3.60 ± 0.60%), but the highest cellulose (58.30 ± 2.06%), and hemicellulose (20.00 ± 3.07%) levels. DP, OP, and PC had around 30% cellulose, and 10% hemicellulose. OP had the highest lignin content (29.00 ± 6.40%), while TW contained (15.70 ± 2.67% cellulose, 13.70 ± 0.002% hemicellulose, and 17.90 ± 4.00% lignin). Among 91 isolated microorganisms, nine were selected for cellulase, xylanase, and/or laccase production. The ability of Bacillus mojavensis to produce laccase and cellulase, as well as B. safensis to produce cellulase and xylanase, is being reported for the first time. In submerged conditions, TW was the most suitable substrate for enzyme production. In this conditions, T. versicolor K1 was the only strain able to produce laccase (4,170 ± 556 U/L). Additionally, Coniocheata hoffmannii P4 exhibited the highest cellulase activity (907.62 ± 26.22 U/L), and B. mojavensis Y3 the highest xylanase activity (612.73 ± 12.73 U/L). T. versicolor K1 culture showed reducing sugars accumulation of 18.87% compared to initial concentrations. Sucrose was the predominant sugar detected by HPLC analysis (13.44 ± 0.02g/L). Our findings suggest that T. versicolor K1 holds promise for laccase production, while TW represents a suitable substrate for sucrose production.

Characterization of a novel multifunctional β-glucosidase/xylanase/feruloyl esterase and its effects on improving the quality of Longjing tea

Herein, a novel multifunctional enzyme β-glucosidase/xylanase/feruloyl esterase (GXF) was constructed by fusion of β-glucosidase and bifunctional xylanase/feruloyl esterase. The activities of β-glucosidase, xylanase, feruloyl esterase and acetyl xylan esterase displayed by GXF were 67.18 %, 49.54 %, 38.92 % and 23.54 %, respectively, higher than that of the corresponding single functional enzymes. Moreover, the GXF performed better in enhancing aroma and quality of Longjing tea than the single functional enzymes and their mixtures. After treatment with GXF, the grassy and floral odors of tea infusion were significantly improved. Moreover, GXF treatment could improve concentrations of flavonoid aglycones of myricetin, kaempferol and quercetin by 68.1-, 81.42- and 77.39-fold, respectively. In addition, GXF could accelerate the release of reducing sugars, ferulic acid and xylo-oligosaccharides by 9.48-, 8.25- and 4.11-fold, respectively. This multifunctional enzyme may have potential applications in other fields such as food production and biomass degradation.

Potential Role of Lauric Acid in Milk Fat Synthesis in Chinese Holstein Cows Based on Integrated Analysis of Ruminal Microbiome and Metabolome.

The composition and metabolic profile of the ruminal microbiome have an impact on milk composition. To unravel the ruminal microbiome and metabolome affecting milk fat synthesis in dairy cows, 16S rRNA and internal transcribed spacer (ITS) gene sequencing, as well as ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS/MS) methods were used to investigate the significant differences in ruminal bacterial and fungal communities as well as metabolome among Chinese Holstein cows with contrasting milk fat contents under the same diet (H-MF 5.82 ± 0.41% vs. L-MF 3.60 ± 0.12%). Another objective was to culture bovine mammary epithelial cells (BMECs) to assess the effect of metabolites on lipid metabolism. Results showed that the acetate-to-propionate ratio and xylanase activity in ruminal fluid were both higher in H-MF. Microbiome sequencing identified 10 types of bacteria and four types of fungi differently abundant at the genus level. Metabolomics analysis indicated 11 different ruminal metabolites between the two groups, the majority of which were lipids and organic acids. Among these, lauric acid (LA) was enriched in fatty acid biosynthesis with its concentration in milk fat of H-MF cows being greater (217 vs. 156 mg per 100 g milk), thus, it was selected for an in vitro study with BMECs. Exogenous LA led to a marked increase in intracellular triglyceride (TG) content and lipid droplet formation, and it upregulated the mRNA abundance of fatty acid uptake and activation (CD36 and ACSL1), TG synthesis (DGAT1, DGAT2 and GPAM), and transcriptional regulation (SREBP1) genes. Taken together, the greater relative abundance of xylan-fermenting bacteria and fungi, and lower abundance of bacteria suppressing short-chain fatty acid-producing bacteria or participating in fatty acid hydrogenation altered lipids and organic acids in the rumen of dairy cows. In BMECs, LA altered the expression of genes involved in lipid metabolism in mammary cells, ultimately promoting milk fat synthesis. Thus, it appears that this fatty acid plays a key role in milk fat synthesis.

Heterologous expression and structure prediction of a xylanase identified from a compost metagenomic library

Xylanases are key biocatalysts in the degradation of the β‐1,4‐glycosidic linkages in the xylan backbone of hemicellulose. These enzymes are potentially applied in a wide range of bioprocessing industries under harsh conditions. Metagenomics has emerged as powerful tools for the bioprospection and discovery of interesting bioactive molecules from extreme ecosystems with unique features, such as high temperatures. In this study, an innovative combination of function-driven screening of a compost metagenomic library and automatic extraction of halo areas with in-house MATLAB functions resulted in the identification of a promising clone with xylanase activity (LP4). The LP4 clone proved to be an effective xylanase producer under submerged fermentation conditions. Sequence and phylogenetic analyses revealed that the xylanase, Xyl4, corresponded to an endo-1,4-β-xylanase belonging to glycosyl hydrolase family 10 (GH10). When xyl4 was expressed in Escherichia coli BL21(DE3), the enzyme activity increased about 2-fold compared to the LP4 clone. To get insight on the interaction of the enzyme with the substrate and establish possible strategies to improve its activity, the structure of Xyl4 was predicted, refined, and docked with xylohexaose. Our data unveiled, for the first time, the relevance of the amino acids Glu133 and Glu238 for catalysis, and a close inspection of the catalytic site suggested that the replacement of Phe316 by a bulkier Trp may improve Xyl4 activity. Our current findings contribute to enhancing the catalytic performance of Xyl4 towards industrial applications.Key points• A GH10 endo-1,4-β-xylanase (Xyl4) was isolated from a compost metagenomic library• MATLAB’s in-house functions were developed to identify the xylanase-producing clones• Computational analysis showed that Glu133 and Glu238 are crucial residues for catalysisGraphical abstract

Unveiling a Microexon Switch: Novel Regulation of the Activities of Sugar Assimilation and Plant-Cell-Wall-Degrading Xylanases and Cellulases by Xlr2 in Trichoderma virens.

Functional microexons have not previously been described in filamentous fungi. Here, we describe a novel mechanism of transcriptional regulation in Trichoderma requiring the inclusion of a microexon from the Xlr2 gene. In low-glucose environments, a long mRNA including the microexon encodes a protein with a GAL4-like DNA-binding domain (Xlr2-α), whereas in high-glucose environments, a short mRNA that is produced encodes a protein lacking this DNA-binding domain (Xlr2-β). Interestingly, the protein isoforms differ in their impact on cellulase and xylanase activity. Deleting the Xlr2 gene reduced both xylanase and cellulase activity and growth on different carbon sources, such as carboxymethylcellulose, xylan, glucose, and arabinose. The overexpression of either Xlr2-α or Xlr2-β in T. virens showed that the short isoform (Xlr2-β) caused higher xylanase activity than the wild types or the long isoform (Xlr2-α). Conversely, cellulase activity did not increase when overexpressing Xlr2-β but was increased with the overexpression of Xlr2-α. This is the first report of a novel transcriptional regulation mechanism of plant-cell-wall-degrading enzyme activity in T. virens. This involves the differential expression of a microexon from a gene encoding a transcriptional regulator.

Sugarcane bagasse derived xylooligosaccharides produced by an arabinofuranosidase/xylobiohydrolase from Bifidobacterium longum in synergism with xylanases

Arabinoxylan is a major hemicellulose in the sugarcane plant cell wall with arabinose decorations that impose steric restrictions on the activity of xylanases against this substrate. Enzymatic removal of the decorations by arabinofuranosidases can allow a more efficient arabinoxylan degradation by xylanases. Here we produced and characterized a recombinant Bifidobacterium longum arabinofuranosidase from glycoside hydrolase family 43 (BlAbf43) and applied it, together with GH10 and GH11 xylanases, to produce xylooligosaccharides (XOS) from wheat arabinoxylan and alkali pretreated sugarcane bagasse. The enzyme synergistically enhanced XOS production by GH10 and GH11 xylanases, being particularly efficient in combination with the latter family of enzymes, with a degree of synergism of 1.7. We also demonstrated that the enzyme is capable of not only removing arabinose decorations from the arabinoxylan and from the non-reducing end of the oligomeric substrates, but also hydrolyzing the xylan backbone yielding mostly xylobiose and xylose in particular cases. Structural studies of BlAbf43 shed light on the molecular basis of the substrate recognition and allowed hypothesizing on the structural reasons of its multifunctionality.

Activities of cell‐wall‐degrading enzymes produced by Pestalotiopsis guepinii

AbstractPestalotiopsis guepinii is a pathogenic fungus that causes grey blight on Camellia pitardii. In this study, we investigated the enzyme activity and kinetics of these cell‐wall‐degrading enzymes (CWDEs) produced by Pestalotiopsis guepinii in both C. pitardii leaves and culture medium. Our enzyme activity experiments revealed that the activities of xylanase, pectin methyl‐galacturonase (PMG), β‐1,4‐endoglucanase (Cx), and β‐glucosidase were high in both C. pitardii leaves and culture medium. These enzymes played a significant role in the pathogenic process. However, the activity of laccase was found to be very low and had a minor impact on the pathogenic process. Furthermore, our enzyme dynamics experiments demonstrated that the optimal reaction temperature for PMG and Cx was 50°C, while for β‐glucosidase and xylanase, it was 60°C. The optimal reaction pH for Cx, β‐glucosidase, and xylanase was 5.0, whereas for PMG, it ranged from 5.0 to 6.0. This indicates that these four enzymes prefer acidic conditions. Moreover, we observed that the activities of Cx, PMG, and xylanase decreased with increasing reaction time. On the other hand, the activity of β‐glucosidase initially increased sharply and then decreased slowly. The maximum reaction rates of the four cell‐wall‐degrading enzymes were ranked as follows: xylanase > PMG > β‐glucosidase > Cx. Additionally, the affinities of these enzymes with substrates were ranked as follows: PMG < Cx < xylanase < β‐glucosidase.

Effectiveness mutagenesis of Bacillus subtilis using EMS (Ethyl Methane Sulfonate) to increase xylanase enzyme activity

This research was conducted to determine the effect of mutagenizing Bacillus subtilis with Ethyl Methane Sulfonate (EMS) for xylanase production and evaluate the effect of different xylan concentrations from corn cobs. The xylanase enzyme is an enzyme that can reduce the xylan content which is an anti-nutrient in animal feed. The wild-type Bacillus subtilis was treated with 50 μg/ml and 100 μg/ml of EMS. The mutants generated were selected for xylanase production in a medium containing xylan from corn cobs as a carbon source. The parameters observed included: Total Plate Count (TPC) and reducing sugar (xylose). Protein concentration and xylanase enzymes were analyzed using the Bradford method and 3.5 Dinitro salicylic acid for reducing sugar according to Miller methods respectively. Five mutants developed from each of the EMS concentrations. Approximately 6 and 6.8 % of the mutants developed from 50 μg/ml and 100 μg/ml of EMS had higher xylanase activities than the wild type and protein and xylose concentrations were higher than the wild type. From the results of the research, it can be seen that the use of EMS compounds has the potential to increase enzyme activity so that it can be potential in reducing antinutrients in animal feedstuff.

Enhancing Xylanase Production from Aspergillus tamarii Kita and Its Application in the Bioconversion of Agro-Industrial Residues into Fermentable Sugars Using Factorial Design

The endo-1,4-β-xylanases (EC 3.2.1.8) are the largest group of hydrolytic enzymes that degrade xylan, the major component of hemicelluloses, by catalyzing the hydrolysis of glycosidic bonds β-1,4 in this polymer, releasing xylooligosaccharides of different sizes. Xylanases have considerable potential in producing bread, animal feed, food, beverages, xylitol, and bioethanol. The fungus Aspergillus tamarii Kita produced xylanases in Adams’ media supplemented with barley bagasse (brewer’s spent grains), a by-product from brewery industries. The culture extract exhibited two xylanase activities in the zymogram, identified by mass spectrometry as glycosyl hydrolase (GH) families 10 and 11 (GH 10 and GH 11). The central composite design (CCD) showed excellent predictive capacity for xylanase production (23.083 U mL−1). Additionally, other enzyme activities took place during the submerged fermentation. Moreover, enzymatic saccharification based on a mixture design (MD) of three different lignocellulosic residues was helpful in the production of fermentable sugars by the A. tamarii Kita crude extract.