Shu-Juan Gao and Jun-Qing Wang contributed equally to this work.
Engineering hyperthermostability into a mesophilic family 11 xylanase from Aspergillus oryzae by in silico design of N-terminus substitution†
Article first published online: 1 NOV 2012
Copyright © 2012 Wiley Periodicals, Inc.
Biotechnology and Bioengineering
Volume 110, Issue 4, pages 1028–1038, April 2013
How to Cite
Gao, S.-J., Wang, J.-Q., Wu, M.-C., Zhang, H.-M., Yin, X. and Li, J.-F. (2013), Engineering hyperthermostability into a mesophilic family 11 xylanase from Aspergillus oryzae by in silico design of N-terminus substitution. Biotechnol. Bioeng., 110: 1028–1038. doi: 10.1002/bit.24768
- Issue published online: 22 FEB 2013
- Article first published online: 1 NOV 2012
- Accepted manuscript online: 23 OCT 2012 01:21PM EST
- Manuscript Accepted: 11 OCT 2012
- Manuscript Revised: 6 OCT 2012
- Manuscript Received: 5 MAY 2012
- National Nature Science Foundation of China. Grant Number: 31101229
- Fundamental Research Funds for the Central Universities of China. Grant Number: JUDCF11032
- Aspergillus oryzae;
- N-terminus substitution;
- in silico design
A mesophilic xylanase from Aspergillus oryzae CICC40186 (abbreviated to AoXyn11A) belongs to glycoside hydrolase family 11. The thermostability of AoXyn11A was significantly improved by substituting its N-terminus with the corresponding region of a hyperthermostable family 11 xylanase, EvXyn11TS. The suitable N-terminus of AoXyn11A to be replaced was selected by the comparison of B-factors between AoXyn11A and EvXyn11TS, which were generated and calculated after a 15 ns molecular dynamic (MD) simulation process. Then, the predicted hybrid xylanase (designated AEx11A) was modeled, and subjected to a 2 ns MD simulation process for calculating its total energy value. The N-terminus substitution was confirmed by comparing the total energy value of AEx11A with that of AoXyn11A. Based on the in silico design, the AEx11A was constructed and expressed in Pichia pastoris GS115. After 72 h of methanol induction, the recombinant AEx11A (reAEx11A) activity reached 82.2 U/mL. The apparent temperature optimum of reAEx11A was 80°C, much higher than that of reAoXyn11A. Its half-life was 197-fold longer than that of reAoXyn11A at 70°C. Compared with reAoXyn11A, the reAEx11A displayed a slight alteration in Km but a decrease in Vmax. Biotechnol. Bioeng. 2013; 110: 1028–1038. © 2012 Wiley Periodicals, Inc.