啤酒泡沫稳定性是评估啤酒质量的一个关键因素,发酵液中的蛋白酶A会影响啤酒的泡沫稳定性。通过过表达钙转运蛋白Gdt1促进蛋白酶A的液泡分选的方法,达到减弱酿酒酵母向细胞外分泌蛋白酶A的目的。以酿酒酵母菌株W303-1A为亲本菌株,通过同源重组依赖型DNA装配方法构建Gdt1过表达菌株,然后将Gdt1过表达菌株和亲本菌株在相同的低氮胁迫条件下进行发酵,测定其胞内外的蛋白酶A活力以及基本发酵性能。研究结果表明:发酵结束时,Gdt1过表达菌株的胞内蛋白酶A活力较亲本菌株提高了12.59%,胞外蛋白酶A活力较亲本菌株降低了18.81%;同时,过表达Gdt1没有影响菌株的基本发酵性能。研究揭示了一种有效的降低蛋白酶A的胞外分泌的方法,为筛选低蛋白酶A胞外分泌菌株提供了理论参考。
Beer foam stability, a key factor in assessing overall beer quality, is affected by proteinase A (PrA) in the fermentation broth. To weaken PrA excretion from Saccharomyces cerevisiae, the vacuolar sorting of PrA was strengthened by overexpression of Ca2+ transporter Gdt1. S. cerevisiae W303-1A was used as the parental strain, and Gdt1-overexpression strain was constructed by exploiting the in vivo homologous recombination-dependent DNA assembler. Fermentation experiments for the Gdt1-overexpression strain and the parental strain were performed under nitrogen-deficient condition, and PrA activity, and fermentation performance of the two strains were examined. Results showed that the intracellular PrA activities of the Gdt1-overexpression strain were increased by 12.59% and the extracellular PrA activities were decreased by 18.81% at the end of fermentation compared with those of the parental strain. Meanwhile, no obvious differences were found on the fermentation performance of the two strains. The research uncovers an effective method to decrease the extracellular secretion of S. cerevisiae PrA, which provides a theoretical reference for the screening of low-PrA-activity strains.
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