Purification and characterization of a thermostable xylanase from the brown-rot fungus Laetiporus sulphureus(ENZYMOLOGY, PROTEIN ENGINEERING, AND ENZYME TECHNOLOGY)

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A thermostable extracellular xylanase was purified and characterized from brown-rot basidiomycete Laetiporus sulphureus, cultivated on biologically pretreated Pinus densiflora biomass. After three consecutive purification steps using DEAE, Mono Q, and Superdex 75 columns, the xylanase specific activity was found to be 72.4U/mg, nine fold higher than that of the crude culture solution, purity was 96%, and the molecular mass determined to be 69.3kDa. The optimal pH and temperature for xylanase activity were 3.0 and 80℃, respectively. Although activity of xylanase was highest at 80℃, it showed highest thermostability at 60℃, retaining approximately 97% of its relative activity following incubation for 4h. In the presence of 5mM solution of CaCl_2, the relative xylanase activity increased by 35.9%; however, it decreased significantly in the presence of 10mM solution of Cu^<2+>. Among the xylan-based substrates tested, purified L. sulphureus xylanase showed the highest activity on beechwood xylan. Thin-layer chromatography (TLC) experiments revealed that purified L. sulphureus xylanase is an endoxylanase that hydrolyzes xylotriose, xylotetraose, and xylopentaose but not xylobiose.
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