Metabolic switching of sake yeast by kimoto lactic acid bacteria through the [GAR+] non-genetic element

Abstract

In traditional kimoto-type sake production, cells of Saccharomyces cerevisiae sake yeast are grown in a starter mash generated by lactate fermentation by lactic acid bacteria (LAB) such as Leuconostoc mesenteroides and Lactobacillus sakei. However, the microbial interactions between sake yeast and kimoto LAB have not been well analyzed. Since the formation of a prion-like element (designated [GAR⁺]) in yeast cells is promoted by bacteria, we here examined the associated phenotype (i.e., increased glucosamine resistance) in sake yeast strains K701 (a representative sake strain) and Km67 (a strain isolated from kimoto-type sake mash). Approximately 0.5% of K701 and Km67 cells, as well as 0.2% of laboratory strain X2180 cells, exhibited increased glucosamine resistance under pure culture conditions, and the frequency of this metabolic switching was further enhanced by coculture with kimoto LAB. The LAB-promoted emergence of the glucosamine-resistant cells was the most prominent in Km67, suggesting that this strain possesses an advanced mechanism for response to LAB. While the glucosamine-resistant clones of X2180 and K701 exhibited lower rates of alcoholic fermentation under high-glucose conditions than did the respective naive strains, glucosamine resistance did not severely affect alcoholic fermentation in Km67. The population of dead cells after alcoholic fermentation was decreased in the glucosamine-resistant clones of X2180, K701, and Km67. These results suggested that the formation of [GAR⁺] in Km67 may be beneficial in kimoto-type sake making, since [GAR⁺] may increase cell viability in the sake starter mash without impairing alcoholic fermentation performance.

Keywords: Alcoholic fermentation; Kimoto lactic acid bacteria; Lactobacillus sakei; Leuconostoc mesenteroides; Saccharomyces cerevisiae; Sake yeast; Stress tolerance; [GAR(+)].