Directed evolution of maltogenic amylase from Bacillus licheniformis R-53: Enhancing activity and thermostability improves bread quality and extends shelf life

Maltogenic amylase from Bacillus licheniformis R-53 improves bread quality and shelf life. Here, we constructed a random mutation library of this enzyme using error-prone PCR to identify mutants with higher activity and thermostability, then screened the key point mutations. Mutant V296F/K418I had 2.16 times the specific activity of the wild-type enzyme, and its temperature for optimum activity increased from 60 to 65 °C. Moreover, it maintained > 60% residual activity after heating at 55 to 80 °C for 30 min. Mixolab experiments showed that treatment with 45 ppm V296F/K418I mutant maltogenic amylase had better effects in delaying dough recrystallization than 60 ppm wild-type enzyme. During bread storage, 45 ppm V296F/K418I was more effective in reducing hardness, improving elasticity, and maintaining sensory than 60 ppm wild-type enzyme. Directed evolution of maltogenic amylase significantly improved its activity and thermostability; the mutant enzyme is conducive to improving bread quality and extending its shelf life.