Characterization of a GH Family 43 β-Xylosidase Having a Novel Carbohydrate-binding Module from Strain TW1.

strain TW1, a gram-positive facultative anaerobic bacterium, was isolated as a xylanolytic microorganism from the wastes of a pineapple processing factory. A gene encoding one of its xylanolytic enzymes, a β-xylosidase, was cloned and sequenced. Sequence analysis revealed that this β-xylosidase, named Xyl43A, was composed of a glycoside hydrolase (GH) family 43 subfamily 12 catalytic module and an unknown function module (UM). The full-length Xyl43A (Xyl43A) was heterologously expressed in and purified. Recombinant Xyl43A exhibited hydrolysis activity against both -nitrophenyl-β-D-xylopyranoside (NPX) and -nitrophenyl-α-L-arabinofuranoside at specific activities of 250 and 310 mU/mg, respectively. The optimal reaction pH and temperature for NPX hydrolysis were 7.1 and 54 ˚C, respectively. At pH 7.0 and 54 ˚C, the and for NPX were 1.2 mM and 2.8 ± 0.15 s, respectively. It was also discovered that the recombinant unknown function module of Xyl43A (Xyl43A-UM) could bind to insoluble xylans like birchwood xylan and oat spelt xylan, whereas it did not bind to cellulosic substrates such as ball-milled cellulose, carboxymethyl cellulose or lichenan. The Xyl43A-UM's binding constant value for oat spelt xylan was 2.0 × 10 M. These results suggest that Xyl43A possesses a novel carbohydrate-binding module, named as CBM91, specific for xylan-containing polysaccharides.