Isolation of sake yeast strains possessing various levels of succinate- and/or malate-producing abilities by gene disruption or mutation

Succinate and malate are the main taste components produced by yeast during sake (Japanese alcohol beverage) fermentation. Sake yeast strains possessi ng various organic acid productivities were isolated by gene disruption. Sa ke fermented using the aconitase gene (ACO1) disruptant contained a two-fol d higher concentration of malate and a two-fold lower concentration of succ inate than that made using the wild-type strain K901. The fumarate reductas e gene (OSM1) disruptant produced sake containing a 1.5-fold higher concent ration of succinate as compared to the wild-type, whereas the a-ketoglutara te dehydrogenase gene (KGD1) and fumarase gene (FUM1) disruptants gave lowe r succinate concentrations. The Delta kgd1 disruptant exhibited lower succi nate productivity in the earlier part of the sake fermentation, while the D elta fum1 disruptant showed lower succinate productivity later in the ferme ntation, indicating that succinate is mainly produced by an oxidative pathw ay of the TCA cycle in the early phase of sake fermentation and by a reduct ive pathway in the later phases. Sake yeasts with low succinate productivit y and/or high malate productivity was bred by isolating mutants unable to a ssimilate glycerol as a carbon source. Low malate-producing yeasts were als o obtained from phenyl succinate-resistant mutants. The mutation of one of these mutant strains with low succinate productivity was found to occur in the KGD1 gene. These strains possessing various succinate- and/or malate-pr oducing abilities are promising for the production of sake with distinctive tastes.