2. Academic Validation
  3. Engineering of isoamylase: improvement of protein stability and catalytic efficiency through semi-rational design

Engineering of isoamylase: improvement of protein stability and catalytic efficiency through semi-rational design

  • J Ind Microbiol Biotechnol. 2016 Jan;43(1):3-12. doi: 10.1007/s10295-015-1708-4.
Youran Li 1 2,  Liang Zhang 3 4,  Zhongyang Ding 5 6,  Zhenghua Gu 7 8,  Guiyang Shi 9 10
Affiliations

Affiliations

  • 1 Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China. liyouran@jiangnan.edu.cn.
  • 2 National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, People's Republic of China. liyouran@jiangnan.edu.cn.
  • 3 Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China. zhangl@jiangnan.edu.cn.
  • 4 National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, People's Republic of China. zhangl@jiangnan.edu.cn.
  • 5 Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China. bioding_jnu@yahoo.com.
  • 6 National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, People's Republic of China. bioding_jnu@yahoo.com.
  • 7 Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China. 327984744@qq.com.
  • 8 National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, People's Republic of China. 327984744@qq.com.
  • 9 Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China. gyshi@jiangnan.edu.cn.
  • 10 National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, People's Republic of China. gyshi@jiangnan.edu.cn.
PMID: 26597030 DOI: 10.1007/s10295-015-1708-4
Abstract

Isoamylase catalyzes the hydrolysis of α-1,6-glycosidic linkages in glycogen, amylopectin and α/β-limit dextrins. A semi-rational design strategy was performed to improve catalytic properties of isoamylase from Bacillus lentus. Three residues in vicinity of the essential residues, Arg505, Asn513, and Gly608, were chosen as the mutation sites and were substituted by Ala, Pro, Glu, and Lys, respectively. Thermal stability of the mutant R505P and acidic stability of the mutant R505E were enhanced. The k cat /K m values of the mutant G608V have been promoted by 49%, and the specific activity increased by 33%. This work provides an effective strategy for improving the catalytic activity and stability of isoamylase, and the results obtained here may be useful for the improvement of catalytic properties of other α/β barrel enzymes.

Keywords

Bacillus lentus; Catalytic efficiency; Isoamylase; Semi-rational design; Structure model.

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