Cloning, sequencing, and expression of the gene encoding an intracellular beta-D-xylosidase from Streptomyces thermoviolaceus OPC-520

The intracellular beta -xylosidase was induced when Streptomyces thermoviol aceus OPC-520 was grown at 50 degreesC in a minimal medium containing xylan or xylooligosaccharides. The 82-kDa protein with beta -xylosidase activity was partially purified and its N-terminal amino acid sequence was analyzed . The gene encoding the enzyme was cloned, sequenced, and expressed in Esch erichia coli. The bxlA gene consists of a 2,100-by open reading frame encod ing 770 amino acids. The deduced amino acid sequence of the bxlA gene produ ct had significant similarity with beta -xylosidases classified into family 3 of glycosyl hydrolases. The bxlA gene was expressed in E. coli, and the recombinant protein was purified to homogeneity. The enzyme was a monomer w ith a molecular mass of 82 kDa.. The purified enzyme showed hydrolytic acti vity towards only p-nitrophenyl-beta -D-xylopyranoside among the synthetic glycosides tested. Thin-layer chromatography analysis showed that the enzym e is an exo-type enzyme that hydrolyze xylooligosaccharides, but had no act ivity toward xylan. High activity against pNPX occurred in the pH range 6.0 -7.0 and temperature range 40-50 degreesC.