Induction of production and secretion β(1→4) glucanase with Saccharomyces cerevesiae by replacing the MET10 gene with egl1 gene from Trichoderma reesei

Aims:  To construct novel brewer's yeast strains with the ability to degrade β-glucan and increase sulfite levels in beer brewing by genetic manipulation.
Methods and Results:  The recombinant plasmid pA15ME containing P_met10-egl1-T_met10 expression cassette was constructed. Bam HI-linearized target plasmid pA15ME was transformed into the industrial brewer's yeast strain Z0103 to replace the MET10 locus through one-step gene replacement. The recombinants Z8, Z7 and Z3 with the ability to secrete active endo-β-1,4-glucanase I into the culture medium were isolated by Congo red dyeing. The enzymatic activities of EG I of Z8, Z7 and Z3 were 3·3, 1·5, 1·3 U l^?1, and the hydrolysing degrees of β-glucans in wort were increased 11·9%, 8·6% and 6·9%, respectively, than that of original strain Z0103. The MET10 gene deletions were confirmed by real-time PCR, and the sulfite levels of the culture mediums inoculated with Z8, Z7 and Z3 were increased 26%, 16% and 17%, respectively, compared to that of Z0103.
Conclusions:  The novel endoglucanase-producing brewer's yeast strains with inserted endoglucanase gene and deficient MET10 gene led to reduced content of barley β-glucans, enhanced filterability and increased sulfur dioxide in fermenting wort. Thus, the cost for addition of microbial β-glucanase enzyme and sulfite preparations in normal beer brewing processes could be reduced.
Significance and Impact of the Study:  These results suggested that genetic engineering approach is a powerful tool to construct the novel recombinant brewer's yeast strains with different properties to reduce the cost of beer brewing and improve the flavour of a beer, and the strains obtained have potential application value in beer brewing.