Screening Of Acidophilic Xylanase High-yield Strain And The Research On Its Characterization And Application

Xylanase can be present in different characterization depending on its source.Due to its complex characterization, xylanase can be applicatied in many industries.Acidophilic xylanase retains high xylanase activity even in low pH, and it shows greatprotential application in clarification of juices, improving nutritional quality of silage andgreenfeed, wine-making industry and the other fields. So far, however, lacking ofxylanase produced strains、the cost of enzyme production coupled with low enzymeyields are the major challenges in industrial applications of acidophilic xylanse. Strategies towards overcoming these challenges have mainly focused on hyper-producingmicrobial strains and the use of cheaper substrates.In this study, an acidophilic xylanase producing strain XAF01was isolatedfrom soil sample collected from shandong povince, and it was identified as Peniciliumjanthinellum， based on morphological, biochemical and physiological tests, and bymean of18SrDNA sequencing. Strain XAF01can produce acidophilic xylanase usingcorncob as the only carbon source. The influence of several factors on xylanaseproduction from Penicillium janthinellium XAF01was evaluated by liquid-statefermention. Maximum xylanase activity was observed in the presence of6%corncob and1.5%ammonium sulphate as the most appropriate inorganic nitrogen source. And themaximum xylanase activity of1807.9U/mL,3.6-fold increase of extracellular xylanaseactivity was obtained after cultivating for7days under optimised conditions. The finalpH of fermention liquid was as low as1.3~2.1.Three extracellular xylanases（XynA、XynB、XynC）were separated byammonium sulfate precipitation and SP-SephroseTMFast Flow cation exchangechromatography. Finally, one of the three extracellular xylanase (XynA) was purified to6.3fold with a50.1%yield by Q-SepharoseTMFast Flow anion exchangechromatography. The molecular mass of Xyn A was approximately25.2kDa.The optimum pH and temperature for the purified XynA were4.8and50℃,respectively, and XynA was found to be stable over an acidic pH value range(3.8~4.8) at50℃, and70%xylanase activity can be detected after cultiviting for5h at40~50℃.XynA showed high substrate specificity on water-soluble corncob xylan, and xylanaseactivity werer not detected using other polysaccharides nor synthetic glycoside(pNP-β-D-cellobioside, pNP-β-D-xylopyranoside, pNP-α-L-arabinofuranoside, etc.) assubstrates. The Kmand Vmaxvalues of the purified xylanase on beechwood xylan was3.14±0.25mg/mL and2407±69μmol/min/mg, respectively. The purified xylanase produced xylooligosaccharide as the end-produce of birchwood xylan, beechwood xylan,water-solubility corncob xylan and corncob liquid (high-temperature and high-pressuretreated) without xylose detected by TLC.The pure xylanase preparation was used for saccharification of corncob liquidwhich had been treated by high-temperature and high-pressure. The effect of differentparameters like pH, temperature, enzyme dosage, time of reaction were firstly studied bysingle factor experiment and further response surface method was employed to optimizedsaccharification yied. Under optimized condition maximum xylooligosaccharide（X2-X6）and (X2+X3) yield were found to be74.0%,9.45mg/mL and62.3%,7.18mg/mL, respectively, which has been improved320.0%and348.7%, compared tothe control.