Cloning, Expression, and Characterization of Xylanase G2 from Aspergillus oryzae VTCC-F187 in Aspergillus niger VTCC-F017

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 BamHI. 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 Hg2+ and partially by Zn2+, Fe2+, and Ag+, while it was slightly stimulated by K+ and EDTA.

Production of xylanase by Aspergillus niger GIO and Bacillus megaterium through solid-state fermentation

“…Aspergillus and Bacillus species have been reported to produce a wide range of enzymes, especially lignocellulosic-degrading enzymes, which are involved in the breaking down of agricultural wastes [13,16,26]. A. niger GIO and B. megaterium used in this work hydrolysed xylan, with evidence of a clear zone on hyrolysed xylan media flooded with iodine against the dark colour of unhydrolysed xylan, as reported by Shakoori et al [27].…”

“…Aspergillus and Bacillus species have been reported to produce a wide range of enzymes, especially lignocellulosic-degrading enzymes, which are involved in the breaking down of agricultural wastes [13, 16, 26]. A. niger GIO and B.…”

“…[21, 22] and others have been used to produce xylanase. Many parameters, such as temperature, pH, metal ions, detergents and others, affect the activities of xylanase [3, 16].…”

Safe production of Aspergillus terreus xylanase from Ricinus communis: gene identification, molecular docking, characterization, production of xylooligosaccharides, and its biological activities

“…For example, the maximum activity in A. niger was at 50 °C [43], and Trichoderma inhamatum was at 50 °C [53]. However, the yield was still similar to that of Aspergillus at 55 °C [54]. Alternatively, because the effective application of end product results from the degradation of Ricinus communis by xylanase, resultant XOS mixtures exhibit good antioxidant activities, which increase gradually with an increasing concentration to 1000 μg/ ml.…”