African Journal of
Microbiology Research

  • Abbreviation: Afr. J. Microbiol. Res.
  • Language: English
  • ISSN: 1996-0808
  • DOI: 10.5897/AJMR
  • Start Year: 2007
  • Published Articles: 5211

Full Length Research Paper

Enhancement of the thermo-alkali-stability of xylanase II from Aspergillus usamii E001 by site-directed mutagenesis

Chenyan Zhou1*, Mingcai Zhang2, Yongtao Wang2, Weiyun Guo1, Zhenhua Liu1, Yan Wang1, and Wu Wang3
1Department of Life Science and Technology, Xinxiang Medical University, Xinxiang 453003, People's Republic of China. 2The First Affiliated Hospital, Xinxiang Medical University, Weihui 453100, People's Republic of China. 3The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122 People's Republic of China.
Email: [email protected]

  •  Accepted: 21 February 2013
  •  Published: 16 April 2013


Replacing several serine and threonine residues on the Ser/Thr surface of xylanase II (Xyn II) from Aspergillus usamii E001 with arginines effectively increased the thermostability of the enzyme. The substitution of Ser and Thr residues on the Ser/Thr surface of the enzyme with four (ST4) or five arginines (ST5) led to an increase in optimal temperature of the enzymes by 2 and 5°C for the ST4 and ST5, respectively. The modified enzymes ST4 and ST5 showed 75 and 87% of maximal activities after incubated for 15 min at 55°Ccompared to only 31% activity for wild-type enzyme. After incubated for 1 h at 55°C, ST4 and ST5 showed 61 and 77% of maximal activity compared to only 27% activity for wild-type enzyme. In addition, these mutations shifted the pH-dependence activity profile to the alkaline region by 1.0 pH units. Kinetic parameters of the four-arginine-substitution enzyme were maintained at the level of the wild-type enzyme with the Km and Vmaxvalues of 5.39±0.20 mg ml-1 and 1410±67 U mg-1 protein, respectively. The five-arginine-substitution enzyme showed only slight alteration in Km and Vmax with Km of 7.68±1.3 mg ml-1 and Vmax of 1161±75 U mg-1 protein, indicating lower substrate affinity and catalytic rate. The study demonstrate that properly introduced arginine residues on the Ser/Thr surface of xylanase might be very effective in the improvement of enzyme thermostability and alkalistability.


Key words: Aspergillus usamii