Abstract

The objectives of the present study were to purify and characterize xylanase enzyme from the fungus obtained from soil. A total of 40 fungi were isolated from 25 soil samples collected after primary screening on Potato Dextrose Agar. In the secondary screening (malt extract agar, 0.5% birch wood xylan), based on the diameter of the clear zone, the fungus was identified as Aspergillus fumigatus by microbial type culture collection (MTCC), Chandigarh, India and was selected for xylanase enzyme production in solid state fermentation using wheat bran. Xylanase was subjected to a three-step purification scheme involving ammonium sulphate precipitation, gel filtration chromatography and anion exchange chromatography. Purity was verified by running the extracted protein on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and a single band was observed. When compared with the standard wide range protein molecular markers on SDS-PAGE, it was found to have a molecular weight of 43 KDa. The K_m and V_max value of xylanase was 3.12 mg/ml and 2857 μmol/min/mg protein as obtained from a Lineweaver-Burk plot. The optimal temperature and pH was found to be 30 and 10°C, respectively. After 4 h of incubation, enzyme retained 100% activity at 30°C. Xylanase was incubated at various pH levels (2 to 12) for 4 h at 30°C, and the residual activity was measured. More than 65% of the original activity was retained at pH ranging from 4 to 10 after 4 h.

Key words: Xylanase, Aspergillus fumigatus, production, enzyme purification, enzyme characterization, Lineweaver–Burk plot.