Abstract
Two α-L-arabinofuranosidases (BfdABF1 and BfdABF3) and a β-D-xylosidase (BfdXYL2) genes were cloned from Bifidobacterium dentium ATCC 27679, and functionally expressed in E. coli BL21(DE3). BfdABF1 showed the highest activity in 50 mM sodium acetate buffer at pH 5.0 and 25°C. This exo-enzyme could hydrolyze p-nitrophenyl arabinofuranoside, arabino-oligosaccharides (AOS), arabinoxylo-oligosaccharides (AXOS) such as 3(2)-α-L-arabinofuranosyl-xylobiose (A(3)X), and 2(3)-α-Larabinofuranosyl-xylotriose (A(2)XX), whereas hardly hydrolyzed polymeric substrates such as debranched arabinan and arabinoxylans. BfdABF1 is a typical exo-ABF with the higher specific activity on the oligomeric substrates than the polymers. It prefers to α-(1,2)-L-arabinofuranosidic linkages compared to α-(1,3)-linkages. Especially, BfdABF1 could slowly hydrolyze 2(3),3(3)-di-α-L-arabinofuranosyl-xylotriose (A(2+3)XX). Meanwhile, BfdABF3 showed the highest activity in sodium acetate at pH 6.0 and 50°C, and it has the exclusively high activities on AXOS such as A(3)X and A(2)XX. BfdABF3 mainly catalyzes the removal of L-arabinose side chains from various AXOS. BfdXYL2 exhibited the highest activity in sodium citrate at pH 5.0 and 55°C, and it specifically hydrolyzed p-nitrophenyl xylopyranoside and xylo-oligosaccharides (XOS). Also, BfdXYL2 could slowly hydrolyze AOS and AXOS such as A(3)X. Based on the detailed hydrolytic modes of action of three exo-hydrolases (BfdABF1, BfdABF3, and BfdXYL2) from Bf. dentium, their probable roles in the hemiceulloseutilization system of Bf. dentium are proposed in the present study. These intracellular exo-hydrolases can synergistically produce L-arabinose and D-xylose from various AOS, XOS, and AXOS.