Construction of a highly thermostable 1,3-1,4-β-glucanase by combinational mutagenesis and its potential application in the brewing industry

To improve the thermostability and catalytic property of a mesophilic 1,3-1,4-β-glucanase by combinational mutagenesis and to test its effect in congress mashing.

A mutant β-glucanase (rE-BglTO) constructed by combinational mutagenesis showed a 25 °C increase in optimal temperature (to 70 °C) a 19.5 °C rise in T ₅₀ value and a 15.6 °C increase in melting temperature compared to wild-type enzyme. Its half-life values at 60 and 70 °C were 152 and 99 min, which were 370 and 800 % higher than those of wild-type enzyme. Besides, its specific activity and k _cat value were 42,734 U mg^-1 and 189 s^-1 while its stability under acidic conditions was also improved. In flask fermentation, the catalytic activity of rE-BglTO reached 2381 U ml^-1, which was 63 % higher than that of wild-type enzyme. The addition of rE-BglTO in congress mashing decreased the filtration time and viscosity by 21.3 and 9.6 %, respectively.

The mutant β-glucanase showed high catalytic activity and thermostability which indicated that rE-BglTO is a good candidate for application in the brewing industry.