ISSN: 0973-7510

E-ISSN: 2581-690X

Xinran Li , Weijia Zhang, Xi Li, Qian Wu, Hui Xu, Dairong Qiao, Yi Cao and Yu Cao
1Microbiology and Metabolic Engineering Key Laboratory of Sichuan Province, College of Life Science, Sichuan University, Chengdu, Sichuan – 610064, P.R. China.
J Pure Appl Microbiol. 2014;8(4):2535-2542
© The Author(s). 2014
Received: 03/05/2014 | Accepted: 26/06/2014 | Published: 31/08/2014

In order to improve the pH and thermal stability of xylanase, the fusion enzyme gene (xynAB) was constructed by Aspergillus niger SCTCC 400264 xylanase genes (xynA and xynB). Based on Aspergillus niger SCTCC400264 xylanase genes xynA (FJ785738.1) and xynB (FJ772090.1), the fusion expression vector pET-32a-xynAB was constructed with an oligopeptide of seven amino acids (GlyGlyGlySerGlyGlyGly) introduced between XYNA and XYNB. After pET-32a-xynAB transformed into E. coli BL21 competent cells, the fusion enzyme was expressed for activity detection by 3, 5-dinitrosalicylic acid (DNS) method and further characterized. The enzyme activities of xynA, xynB and xynAB were 16.58U/mg, 1201.7U/mg and 47.3U/mg, respectively. There appeared two peaks of the optimum temperature of XYNAB, the first peak value was 50ºC, the same with XYNA and the second one was 35ºC, the same with XYNB. The optimum pH of XYNAB was 4.0, which compromised the properties of two single enzymes whose optimum pH were 3.0 and 5.0, respectively. When incubated at 80 ºC for 5min, it retained 35% of enzyme activity, showing higher thermostability than XYNA but lower than XYNB. XYNAB had a good stability at pH ranging from 2.0 to 10.0. The effect of metal ions on it was basically the same as XYNA, XYNB. The enzyme activity of XYNAB was strongly increased by K+ and Mg2+ but inhibited by Cu2+, Fe3+ and Mn2+. The enhanced difunctional XYNAB fusion enzyme was  gained in this paper. According to the conformation of XYNAB, the gap of XYNB for binding substrate was mostly blocked or covered by the whole XYNA, which resulted in XYNB not binding with substrate completely and so XYNAB showed the whole of the XYNA activity and part of XYNB.


Aspergillus niger, Xylanase, Fusion Enzyme, pH stability, Thermal stability

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