Highly alkali-stable and cellulase-free xylanases from Fusarium sp. 21 and their application in clarification of orange juice

Xylanase is a versatile tool in the food, fiber biobleaching and biofuel industries. Here, to discover new enzyme with special properties, we cloned three xylanases (Xyn11A, Xyn11B, and Xyn11C) by mining the genome of the xylanase producing fungus strain Fusarium sp. 21, biochemically characterized these enzyme and explored their potential application in juice processing. Both Xyn11A and Xyn11B had an optimal pH of 6.0 and optimal temperature of 45 °C, and retained >90% of the residual activity at pH range of 5–10.5 for 24 h. Xyn11C displayed the maximum activity at pH 5.0 and 45 °C and outstanding pH stability with a minimal loss of activity in the pH range of 2.0–10.5. These three xylanases displayed a strong specificity towards beechwood and corncob xylan, with no activity for other substrates. Xyn11A showed much a higher activity against corncob xylan, while Xyn11B and Xyn11C presented higher activities against beechwood xylan. Xyn11A catalyzed the hydrolysis of beechwood xylan with a Km of 4.25 ± 0.29 mg·mL−1 and kcat/Km of 30.34 ± 0.65 mL·s−1·mg−1, while the hydrolysis of corncob xylan had Km and kcat/Km values of 14.73 ± 1.43 mg·mL−1and 26.48 ± 0.11 mL·s−1·mg−1, respectively. Xyn11B and Xyn11C hydrolyzed beechwood xylan with Km of 9.8 ± 0.69 mg·mL−1, and 4.89 ± 0.38 mg·mL−1and kcat/Km values of 45.07 ± 1.66 mL−1·mg−1, and 26.95 ± 0.67 mL·s−1·mg−1, respectively. Beechwood xylan hydrolysates catalyzed by these three xylanases contained xylobiose, xylotriose and xylooligosaccharides (XOS). The clarification of orange juice was improved when treated with these three xylanases. Conclusively, the desirable pH stability and substrate specificity make Xyn11A, Xyn11B and Xyn11C have high potential for application in fiber biobleaching, wine and fruit juice clarification, as well as probiotic XOS production.