Biochemical and catalytic properties of an endoxylanase purified from the culture filtrate of Thermomyces lanuginosus ATCC 46882

An endoxylanase (1,4-β-d-xylan xylanohydrolase, EC 3.2.1.8) from the culture filtrates of T. lanuginosus ATCC 46882 was purified to homogeneity by DEAE–Sepharose and Bio-Gel P-30 column chromatographies. The purified endoxylanase had a specific activity of 888.8 μmol min−1 mg −1 protein and accounted for approximately 30% of the total protein secreted by this fungus. The molecular mass of native (non-denatured) and denatured endoxylanase were 26.3 and 25.7 kD as, respectively. Endoxylanase had a pI of 3.7 and was optimally active between pH 6.0–6.5 and at 75 °C. The enzyme showed >50% of its original activity between pH 5.5–9.0 and at 85 °C. The pH and temperature stability studies revealed that this endoxylanase was almost completely stable between pH 5.0–9.0 and up to 60 °C for 5 h and at pH 10.0 up to 55 °C for 5 h. Thin-layer chromatography (TLC) analysis showed that endoxylanase released mainly xylose (Xyl) and xylobiose (Xyl2) from beechwood 4-O-methyl-d-glucuronoxylan, O-acetyl-4-O-methyl-d-glucuronoxylan and rhodymenan (a β-(1→3)–β(1→4)-xylan). Also, the enzyme released an acidic xylo-oligosaccharide from 4-O-methyl-d-glucuronoxylan, and an isomeric xylotetraose and an isomeric xylopentaose from rhodymenan. The enzyme hydrolysed [1-3H]-xylo-oligosaccharides in an endofashion, but the hydrolysis of [1-3H]-xylotriose appeared to proceed via transglycosylation, since the xylobiose was the predominant product. Endoxylanase was not active on pNPX and pNPC at 40 and 100 mM for up to 6 h, but showed some activity towards pNPX at 100 mM after 20–24 h. The results suggested that the endoxylanase from T. lanuginosus belongs to family 11.