Purification and Properties of an Extracellular β-Xylosidase from Aspergillus japonicus and Sequence Analysis of the Encoding Gene

An extracellular protein exhibiting β-xylosidase activity was purified from the culture filtrate of a filamentous fungus, Aspergillus japonicus strain MU-2, grown on oat spelt xylan. The purified enzyme was a monomeric glycoprotein with an apparent Mr of 113.2 kDa as estimated by SDS–PAGE. β-Xylosidase activity was optimal at pH 4.0 and 70°C. The enzyme also showed β-glucosidase and α-l-arabinofuranosidase activities. The genomic DNA and cDNA encoding this protein were cloned and sequenced. Southern blot analysis indicated that the β-xylosidase gene (xylA) was present as a single copy in the genome. An open reading frame, consisting of 2412 bp, was not interrupted by introns, and it encoded a presumed signal peptide of 17 amino acids and a mature protein of 787 amino acids. The deduced amino acid sequence of the xylA gene product showed a high degree of identity (69%) to the primary structure of the Aspergillus niger β-xylosidase XlnD that belongs to the glycoside hydrolase family 3. Moreover, the xylA gene was functionally expressed in the yeast Pichia pastoris.