| Abstract [eng] |
Cloning, expression and biochemical characterization of the thermostable xylanase from Geobacillus thermodenitrificans strain JK1 Xylan is a major structural polysaccharide in plant cells, and is the second most abundant polysaccharide in nature. Due to its heterogeneity and complexity, the complete hydrolysis of xylan requires a variety of cooperatively acting enzymes. These xylanolytic enzyme systems are quite widespread among prokariotic and eukaryotic microorganisms. Endo-1,4-β-xylanase is considered to play the most important role in xylan degradation. Thermostable xylanases have potential applications in a wide range of industrial processes, including pulp and paper industry, food and animal feeds industry and bioconvertion. In this study a thermophilic JK1 strain producing xylanolytic enzymes was isolated from compost sample. The 16S rRNR and spo0A genes sequence analysis indicated that JK1 strain had the highest similarity with G. thermodenitrificans NG80-2 strain. Based on the xylanase gene sequence from G. thermodenitrificans NG80-2 strain, degenerated primers were designed for amplification of xylanase-coding gene (1224 bp) from JK1 strain. Subsequently, the amplified gene was cloned into expression vector (pET-28c(+)) and the recombinant xylanase with a polyhistidine-tag was designed (50,979 kDa). Biochemical properties of the purified by nickel affinity chromatography recombinant xylanase from G. thermodenitrificans JK1 strain were further characterized. The optimal activity of recombinant enzyme was obtained at 70 °C and pH 6,0. It was shown, that this enzyme is thermostable and cellulase-free. |
