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• W2960748081 abstract "Directed evolution methodologies have been used as promising strategies for improving the catalytic properties of many existing enzymes. In the presented work, this approach was applied to improve the enzyme activity of l-asparaginase I obtained from Bacillus megaterium H-1. After two rounds of error-prone polymerase chain reaction (epPCR) and two generations of sequential DNA shuffling, all of 5 different mutants showed a significant increase in the enzyme activity of l-asparaginase I, ranging from 6.27 to 22.78 IU/mL. Among these mutants, D-9B and DD-12G displayed the relatively high catalytic activity, which were 20.22-fold and 21.33-fold higher than the wild-type enzyme (WT), respectively. Furthermore, the catalytic efficiency (kcat/Km) of D-9B and DD-12G were also improved, which were 132.73 min−1mM−1 and 146.39 min−1mM−1, respectively, in comparison to that of WT (3.39 min−1mM−1). In addition, mutant DD-12G showed tolerance toward wider range of pH values and higher temperatures than its WT counterpart. Homology modeling of above two mutants reflected a reduction of hydrogen bonds and an introduction of flexible residues in the loops near the active catalytic site Thr15. These changes contributed to the flexibility of loops, which may lead to further enhancement in catalytic efficiency. Results also showed that approximately 88.5% (0.978 mg/kg) acrylamide could be removed from mutant DD-12G pre-treated fried potato chips. This study clearly shows that directional evolution methods can indeed be utilized to improve the activity of l-asparaginase, which could also provide research basis for future application in food industry." @default.
• W2960748081 created "2019-07-23" @default.
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• W2960748081 date "2019-12-01" @default.
• W2960748081 modified "2023-10-12" @default.
• W2960748081 title "Improvement of the activity of l-asparaginase I improvement of the catalytic activity of l-asparaginase I from Bacillus megaterium H-1 by in vitro directed evolution" @default.
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• W2960748081 doi "https://doi.org/10.1016/j.jbiosc.2019.06.001" @default.
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