An endoxylanase rapidly hydrolyzes xylan into major product xylobiose via transglycosylation of xylose to xylotriose or xylotetraose.

Here, we proposed an effective strategy to enhance a novel endoxylanase (Taxy11) activity and elucidated an efficient catalysis mechanism to produce xylooligosaccharides (XOSs). Codon optimization and recruitment of natural propeptide in Pichia pastoris resulted in achievement of Taxy11 activity to 1405.65 ±â€¯51.24 U/mL. Analysis of action mode reveals that Taxy11 requires at least three xylose (xylotriose) residues for hydrolysis to yield xylobiose. Results of site-directed mutagenesis indicate that residues Glu119, Glu210, and Asp53 of Taxy11 are key catalytic sites, while Asp203 plays an auxiliary role. The novel mechanism whereby Taxy11 catalyzes conversion of xylan or XOSs into major product xylobiose involves transglycosylation of xylose to xylotriose or xylotetraose as substrate, to form xylotetraose or xylopentaose intermediate, respectively. Taxy11 displayed highly hydrolytic activity toward corncob xylan, producing 50.44 % of xylobiose within 0.5 h. This work provides a cost-effective and sustainable way to produce value-added biomolecules XOSs (xylobiose-enriched) from agricultural waste.

Biochemical characterization of a novel xylanase from Paenibacillus barengoltzii and its application in xylooligosaccharides production from corncobs.

A novel xylanase gene (PbXyn10A) from Paenibacillus barengoltzii was cloned and expressed in Escherichia coli. PbXyn10A had an open reading frame of 3,063 bp, and its deduced amino acid sequence shared the highest identity of 72% with a xylanase from Paenibacillus curdlanolyticus. The recombinant xylanase (PbXyn10A) was purified and biochemically characterized. PbXyn10A was most active at pH 6.5 and 60 °C, respectively. It exhibited strict substrate specificity towards birchwood xylan, beechwood xylan and oat-spelt xylan, with Km values of 2.19, 2.04 and 2.51 mg/mL, respectively. The enzyme hydrolyzed xylan to yield mainly xylooligosaccharides (XOS) with degree of polymerization 2-4. A new strategy for XOS production from corncobs pretreated by steam explosion using acidic electrolyzed water, followed by enzymatic hydrolysis was developed. The highest XOS yield of 75% (based on xylan in raw corncobs) was achieved. This is the first report on a xylanase from Paenibacillus barengoltzii.

Production of xylooligosaccharides by autohydrolysis of hazelnut (Corylus avellana L.) shell.

Hazelnut shell (HS), husk and pruning residues were characterized and evaluated for xylooligosaccharides (XOS) production by autohydrolysis. HS contained the highest amount of xylan and yielded more XOS compared to other hazelnut residues. The temperature and holding time of HS autohydrolysis greatly influenced the composition of the liquor and the remaining solid. The highest XOS yield (62% of the feedstock xylan) was obtained at 190°C and 5min of holding time. At this temperature, 30min of holding time was required to maximize the percentage of XOS with low degree of polymerization. Xylose, acetic acid and furfural concentrations increased with treatment severity. The concentrations of the products in the autohydrolysis liquors followed specific trends with changing severity factor (log Ro) values. Solubilization of xylan in the treatments enhanced the cellulose and lignin contents in the remaining solids.

Miscanthus×giganteus xylooligosaccharides: Purification and fermentation.

A procedure was developed to recover xylooligosaccharides (XOS) from Miscanthus×giganteus (M×G) hydrolyzate. M×G hydrolyzate was prepared using autohydrolysis, and XOS rich fractions were acquired using activated carbon adsorption and stepwise ethanol elution. The combined XOS fractions were purified using a series of ion exchange resin treatments. The end product, M×G XOS, had 89.1% (w/w) total substituted oligosaccharides (TSOS) composed of arabinose, glucose, xylose and acetyl group. Bifidobacterium adolescentis and Bifidobacterium catenulatum (health promoting bacteria) were cultured in vitro on M×G XOS and a commercial XOS source, which was used as a comparison. B. adolescentis grew to a higher cell density than B. catenulatum in both XOS cultures. Total xylose consumption for B. adolescentis was 84.1 and 84.8%, respectively for M×G and commercial XOS cultures; and for B. catenulatum was 76.6 and 73.6%, respectively. The xylobiose (X2), xylotriose (X3) and xylotetraose (X4) were almost utilized for both strains. Acetic and lactic acids were the major fermentation products of the XOS cultures.