Purification, biochemical characterization, and structure of recombinant endo-1,4-β-xylanase XylE

The gene xylE encoding endo-1,4-beta-xylanase from the 10th family of glycosyl hydrolases produced by the mycelial fungus Penicillium canescens has been expressed under the control of the strong promoter of the bgaS gene encoding beta-galactosidase from P. canescens. As a result, a strain-producer of endoxylanase XylE was developed. The recombinant enzyme was isolated and purified to homogeneity with specific activity of 50 U/mg. The physicochemical and biochemical properties of the endoxylanase were studied. The maximal enzymatic activity was observed at pH 6.0 and 70A degrees C. Endoxylanase XylE was shown to be a highly thermostable enzyme with half-inactivation period tau(1/2) of 7 h at 60A degrees C. The kinetic parameters were 0.52 mg/ml (K (m)) and 75 mu mol/min per mg (V (max)) using birch xylan as the substrate. Crystals of endoxylonase XylE were obtained, and the 3D structure was solved at 1.47 resolution. The 3D structure of an endo-1,4-beta-xylanase from the 10th family containing carbohydrate and unique cyclic structure located at the C-terminus of the polypeptide chain was obtained for the first time.