Abstract

A non-cellulosomal xylanase from Clostridium thermocellum, XynX, consists of a family-22 carbohydratebinding module (CBM22), a family-10 glycoside hydrolase (GH10) catalytic module, two family-9 carbohydrate-binding modules (CBM9-I and CBM9-II), and an S-layer homology (SLH) module. E. coli BL21(DE3) (pKM29), a transformant carrying xynX', produced several truncated forms of the enzyme. Among them, three major active species were purified by SDS-PAGE, activity staining, gel-slicing, and diffusion from the gel. The truncated xylanases were different from each other only in their C-terminal regions. In addition to the CBM22 and GH10 catalytic modules, XynX1 had the CBM9-I and most of the CBM9-II, XynX2 had the CBM9-I and about 40% of the CBM9-II, and XynX3 had about 75% of the CBM9-I. The truncated xylanases showed higher binding capacities toward Avicel than those toward insoluble xylan. XynX1 showed a higher affinity toward Avicel (70.5%) than XynX2 (46.0%) and XynX3 (42.1%); however, there were no significant differences in the affinities toward insoluble xylan. It is suggested that the CBM9 repeat, especially CBM9-II, of XynX plays a role in xylan degradation in nature by strengthening cellulose binding rather than by enhancing xylan binding.

• Keywords: Clostridium thermocellum XynX; truncated xylanases; gel-elution; carbohydrate-binding module; cellulose and xylan binding