2Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, ul. Vavilova 32, 119991 Moscow, Russia; fax: (499) 135-1405; E-mail: firstname.lastname@example.org
3European Molecular Biology Laboratory (EMBL), c/o DESY, Build. 25A, Notkestrasse 85, 22603 Hamburg, Germany; fax: +49(0)4089-902149; E-mail: email@example.com
* To whom correspondence should be addressed.
Received June 20, 2012; Revision received July 7, 2012
The gene xylE encoding endo-1,4-β-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 β-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 70°C. Endoxylanase XylE was shown to be a highly thermostable enzyme with half-inactivation period τ1/2 of 7 h at 60ºC. The kinetic parameters were 0.52 mg/ml (Km) and 75 µmol/min per mg (Vmax) using birch xylan as the substrate. Crystals of endoxylonase XylE were obtained, and the 3D structure was solved at 1.47 Å resolution. The 3D structure of an endo-1,4-β-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.
KEY WORDS: Penicillium canescens, xylanase, homologous expression, XylE, 3D structure, biochemical and physicochemical properties, substrate specificity