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Journal of Zhejiang University-SCIENCE A

, Volume 7, Issue 11, pp 1948–1955 | Cite as

Enhancement of the thermostability of β-1,3-1,4-glucanase by directed evolution

  • Zhang Xiu-yan 
  • Ruan Hui 
  • Mu Lin 
  • He Guo-qing 
  • Tang Xing-jun 
  • Chen Qi-he 
Article

Abstract

In order to improve the thermostability of β-1,3-1,4-glucanase, evolutionary molecular engineering was used to evolve the β-1,3-1,4-glucanase from Bacillus subtilis ZJF-1A5. The process involves random mutation by error-prone PCR and DNA shuffling followed by screening on the filter-based assay. Two mutants, EGs1 and EGs2, were found to have four and five amino acid substitutions, respectively. These substitutions resulted in an increase in melting temperature from Tm=62.5 °C for the wild-type enzyme to Tm=65.5 °C for the mutant EGs1 and 67.5 °C for the mutant EGs2. However, the two mutated enzymes had opposite approaches to produce reducing sugar from lichenin with either much higher (28%) for the former or much lower (21.6%) for the latter in comparison with their parental enzymes. The results demonstrate that directed evolution is an effective approach to improve the thermostability of a mesophilic enzyme.

Key words

Directed evolution Error-prone PCR DNA shuffling β-1,3-1,4-glucanase Thermostability 

CLC number

Q816 

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Copyright information

© Zhejiang University 2006

Authors and Affiliations

  • Zhang Xiu-yan 
    • 1
  • Ruan Hui 
    • 1
  • Mu Lin 
    • 1
  • He Guo-qing 
    • 1
  • Tang Xing-jun 
    • 1
  • Chen Qi-he 
    • 1
  1. 1.Department of Food Science and NutritionZhejiang UniversityHangzhouChina

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