ABSTRACT
Xylanase is a high-profile glycoside hydrolase with applications in brewing, feed, pharmacy and bioenergy industries, but most of xylanases are in active below 30 ℃. In order to obtain low temperature active xylanase, a xylanase gene, XYN11A, was cloned from Penicillium sp. L1 and expressed in Pichia pastoris GS115. After purification and enzyme assay, optimal pH and temperature were determined to be 3.5 to 4.0 and 55 ℃. This enzyme was stable at acid and neutral condition (pH 1.0 to 7.0) or under the treatment of 40 ℃ for 1 hour. This xylanase displayed strong resistance to all tested ions and chemicals. Noteworthily, XYN11A maintained a higher activity of 6 700 U/mg than a lot of GH11 xylanase, and demonstrated higher activity (24% to 58%) at lower temperature from 20 to 40 ℃. After beechwood xylan hydrolysis for 16 h, the hydrolysates consisted mainly of xylobiose, xylotriose and xylotetraose and barely of xylose, thus XYN11A could be used for the production of prebiotic xylooligosaccharide. Possessing the features of acidophilic, highly active at lower temperature and oligosaccharide production, XYN11A demonstrated great potential in food and feed industrials.
ABSTRACT
Efficient utilization of cellulose and xylan is of importance in the bioethanol industry. In this study, a novel bifunctional xylanase/cellulase gene, Tcxyn10a, was cloned from Thermoascus crustaceus JCM12803, and the gene product was successfully overexpressed in Pichia pastoris GS115. The recombinant protein was then purified and characterized. The pH and temperature optima of TcXyn10A were determined to be 5.0 and 65-70 °C, respectively. The enzyme retained stable under acid to alkaline conditions (pH 3.0-11.0) or after 1-h treatment at 60 °C. The specific activities of TcXyn10A towards beechwood xylan, wheat arabinoxylan, sodium carboxymethyl cellulose and lichenan were (1 480±26) U/mg, (2 055±28) U/mg, (7.4±0.2) U/mg and (10.9±0.4) U/mg, respectively. Homologous modeling and molecular docking analyses indicated that the bifunctional TcXyn10A has a single catalytic domain, in which the substrate xylan and cellulose shared the same binding cleft. This study provides a valuable material for the study of structure and function relationship of bifunctional enzymes.