Cloning, Expression, and Characterization of Xylanase G2 from <i>Aspergillus oryzae</i> VTCC‐F187 in <i>Aspergillus niger</i> VTCC‐F017
Do Thi Tuyen, Nguyễn Tiến Cường, Nguyen Sy Le Thanh, Nguyễn Thị Thảo, Le Thanh Hoang, Nguyen Thi Hien Trang, Nguyễn Thị Trung, Dao Thi Mai Anh
Abstract
The study focuses on engineering of recombinant Aspergillus niger to produce highly active xylanase. The xylanase G2 encoding gene originating from Aspergillus oryzae VTCC‐F187 was cloned, amplified, and inserted into the pAN7.1GluA vector with specific primers possessing Bam HI. The recombinant plasmid was introduced into Aspergillus niger VTCC‐F017 by chemical methods. The recombinant strain was checked by polymerase chain reaction method and Southern blot. Next, the recombinant protein was expressed and purified by His‐tag column. The molecular mass of the purified xylanase G2, as determined by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS‐PAGE), was 21 kDa with a specific activity of 1025 IU/mg towards 0.5% (w/v) of birchwood xylan. The optimal temperature and pH were 55°C and pH 6.5, respectively. The enzyme was stable in a temperature ranges 25–40°C and a pH ranges 5–7. The presence of Tween 80 enhanced xylanase activity. Triton X‐100, however, had no impact on the function of the enzyme. The xylanase activity was reduced by Tween 20, SDS, and organic solvents. The enzyme was completely inhibited by Hg 2+ and partially by Zn 2+ , Fe 2+ , and Ag + , while it was slightly stimulated by K + and EDTA.