Improving Thermostability of β-glucanase from Bacillus subtilis ZJF-1A5 and Analyzing Enzyme Properties of Mutants

In order to obtain β-glucanase with higher thermostablility, directed evolution was used to evolve the β-glucanase gene from Bacillicus subtilis ZJF-1A5, and properties of wild-type and mutated enzyme were analyzed. Two mutants producing thermostable enzymes were obtained, named EGs1 and EGs2, respectively. Compared with the wild-type enzyme, the half-life temperatures of EGs1 and EGs2 enzyme were shifted upward by 3 and 5 ℃, and approached 65.5 and 67.5 ℃, respectively. The optimum temperatures of the two mutated enzymes were increased by 5 ℃, and approached 60 ℃.The abilities to produce reducing sugar from lichenin were either much higher (28%) for the former or much lower (21.6%) for the latter, the affinities of the two mutated enzymes for the lichenin were basically unchanged in comparison with their parental enzymes. The optimum pH was 6.5 for the EGs1 enzyme and wild-type, and 7.0 for EGs2 enzyme; The pH stability for all enzymes was good, the activities of parental enzyme and the two mutated enzymes held level of more than 80% at pH 6.0~8.0 for 48 h, respectively. The results of genetic stability experiments showed that the two mutants had good genetic stability.