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 T m=62.5 °C for the wild-type enzyme to T m=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.
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Project supported by the National Natural Science Foundation of China (No. 20276064) and Natural Science Foundation of Zhejiang Province (No. Z304076), China
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Zhang, Xy., Ruan, H., Mu, L. et al. Enhancement of the thermostability of β-1,3-1,4-glucanase by directed evolution. J. Zhejiang Univ. - Sci. A 7, 1948–1955 (2006). https://doi.org/10.1631/jzus.2006.A1948
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DOI: https://doi.org/10.1631/jzus.2006.A1948