Modification And Expression Of Xylanase (XynⅡ) Gene From Aspergillus Usamii

The xylanase (XynII) from Aspergillus usamii, which was one of the high specific activity xylanase, belonged to family G/11. The xylanase from Thermomonospora fusca belonging to family G/11, could tolerate high temperature. The hybrid xylanase XynIIAB was constructed by the substitution of the N-terminus segment of the Aspergillus usamii xylanase XynII with corresponding region of Thermomonospora fusca xylanase TfxA. The results that the hybrid protein lose it's bioactivity was beyond wanted. And the three-dimensional structures of the proteins were modelinged to compare their difference. One key position (e.g. the sixth Tyrosine in the XynII) was found in the following observation. In the hybrid protein XynIIAB, the key position vanished. Then Y6S mutation was introduced in XynII by site-directed mutagenesis. The mutant xylanase (XynIIY6S) also lost it's bioactivity. The three-dimensional models of other xylanases which belonged to G/11 family were modelinged by Esypred server too. At the same conservative position (e.g. the sixth Tyrosine in XynII), they are all aromatic amino acids(Y/F).XynII was a type of xylanase which had higher relative activity. With S179C-directed mutagenesis, its optimum reaction temperature was improved from 50℃to 52℃. After 70 min of incubation at 50℃, it still had 50% residual activity, while the residual activity of the native xylanase was 50% after only 23 min at the same incubation temperature. The thermostability was enhanced 3 fold. However, there was no obvious change in optimum pH.