P44: Characterization of xylanase from a color variant Aureobasidium pullulans isolated in Thailand

A number of xylanase-producing A. pullulans, both typical pigmented and color variant isolates, were obtained from different leaves of plant collected throughout Thailand. A color variant strain, 109B, was found to exhibit cellulase-free and thermo-activated xylanase activity. The highest xylanase yield (4.10 U.ml^-1) was obtained in the medium containing 1% (w/v) corncob after 3 days of cultivation. The enzyme was purified, using ultrafiltration, ammonium sulphate precipitation (60-80% (w/v)), ion exchange and gel filtration chromatography, and was then characterized. Molecular mass of the enzyme was approximately 72 kDa as determined by SDS-PAGE and the optimal pH and temperature were 6.0 and 70^oC, respectively. The metal ions, Fe²⁺ and Cu²⁺, partially inhibited the enzyme, while Mg²⁺, Co²⁺ and Ca²⁺ slightly enhanced the enzyme activity. Using beech wood xylan as the substrate, the K_m and V_max values of the enzyme were 11.71 mg.ml^-1 and 2.05 µmol.min^-1.mg^-1protein, respectively. This enzyme was relatively stable at 50^oC since it could retain more than half of its original activity after 3-hr incubation period. The thermostability of the enzyme was further improved using polyols (mannitol, sorbitol and glycerol). Addition of mannitol (0.5 mM) to the enzyme solution was revealed to be the best to prolong the half-life of this xylanase around 3.5-fold at 70ºC.