title: Biochemical properties of genetic recombinant xylanase II.
abstract: The aim of this study was to overexpress the xylanase II gene of Trichoderma reesei in Escherichia coli and determine the characteristics of the recombinant enzyme. Recombinant xylanase II gene was constructed by ligating the cDNA of xylanase, obtained from reverse transcriptase-polymerase chain reaction, and fused with NusA protein of pET-431b plasmid. An Ni2+-NTA affinity column was used to further purify the recombinant xylanase II. The molecular mass of the recombinant enzyme measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was approx 76 kDa (including 55 kDa of NusA and 21 kDa of xylanase II), and the isoelectric point and specific activity were 7.5 and 225 U/mg, respectively. The optimal reaction temperature and pH for the recombinant enzyme were 50 degrees C and 4.0, respectively. The recombinant enzyme was stable at a pH range of 5.0-10.0 and maintained 95% residual activity after incubating at 30-35 degrees C for 30 min. The kinetic parameters KM and Vmax of the recombinant xylanase II were 13.8 mg/mL and 336 micromol/(mg.min), respectively, using birchwood xylan as the substrate.
authors.affiliation: Department of Food and Nutrition, Providence University, 200 Chungchi Road, Shalu, Taichung 43301, Taiwan.
authors.forename: Meng-Yun
authors.initials: MY
authors.lastname: Tung
authors.forename: Chen-Tien
authors.initials: CT
authors.lastname: Chang
authors.forename: Yun-Chin
authors.initials: YC
authors.lastname: Chung
mesh_terms: D001483:Base Sequence; D003001:Cloning, Molecular; D043364:Endo-1,4-beta Xylanases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004789:Enzyme Activation; D004795:Enzyme Stability; D004926:Escherichia coli / Q000378:metabolism; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000378:metabolism*; D016415:Sequence Alignment; D013379:Substrate Specificity; D014242:Trichoderma / Q000201:enzymology*
chemical_list: D008670:Metals; D011994:Recombinant Proteins; D043364:Endo-1,4-beta Xylanases