Xylanase B fromClostridium cellulovorans743B: overexpression, purification, crystallization and X-ray diffraction analysis

Abstract

Clostridium cellulovoransproduces multi-enzyme complexes called cellulosomes capable of efficiently degrading cellulosic biomass. There are three xylanase genes containing a sequence corresponding to a dockerin domain that are necessary for constructing cellulosomes in the genome. Among the xylanases encoded by these genes, xylanase B (XynB) contains a catalytic domain belonging to glycoside hydrolase family 10 and a carbohydrate-binding module (CBM) at the N-terminus, making it a member of CBM family 22. In this study, XynB was cloned, overexpressed, purified and crystallized. XynB was crystallized using the hanging-drop vapour-diffusion method in the presence of 0.2 Msodium acetate trihydrate, 0.1 MTris–HCl pH 8.5, 32%(w/v) PEG 4000 at 293 K. X-ray diffraction analysis revealed that the crystal diffracted to 1.95 Å resolution and belonged to space groupP212121, with unit-cell parametersa = 74.28,b= 77.55,c= 88.20 Å, α = β = γ = 90°. The data-evaluation statistics revealed high quality of the collected data, thereby establishing a solid basis for determination of the structure of cellulosomal xylanase fromC. cellulovorans.