Debranching of arabinoxylan: properties of the thermoactive recombinant alpha-L-arabinofuranosidase from Clostridium stercorarium (ArfB)

Abstract

The gene arfB encoding alpha-L-arabinofuranosidase B of the cellulolytic thermophile Clostridium stercorarium was expressed in Escherichia coli from a 2.2-kb EcoRI DNA fragment. The recombinant gene product ArfB was purified by fast-performance liquid chromatography. It has a tetrameric structure with a monomeric relative molecular mass of 5200. The optima for temperature and pH are 70 degrees C and 5.0 respectively. The enzyme appears to have no metal cofactor requirement and is sensitive to sulfhydryl reagents. It hydrolyzes aryl and alkyl alpha-L-arabinofuranosides and cleaves arabinosyl side-chains from arabinoxylan (oat-spelt xylan) and from xylooligosaccharides produced by recombinant endoxylanase XynA from the same organism. The identify of the N-terminal amino acid sequences indicates that ArfB corresponds to the major alpha-arabinosidase activity present in the culture supernatant of C. stercorarium.