Construction of sake yeast with low production of dimethyl trisulfide (DMTS) precursor by a self-cloning method

摘要

Dimethyl trisulfide (DMTS) is the primary component responsible for "hineka", the stale aroma of Japanese sake. Deletion of the MRU or MDI1 gene of sake yeast, encoding 5 '-methylthioribose-1-phosphate isomerase and 5，-meth-ylthioribulose-1-phosphate dehydratase, respectively, has been reported to greatly reduce the amount of DMTS precursor (DMTS-P1) in sake and to suppress the formation of DMTS during storage. In this study, we constructed sake yeast strains lacking MRU gene function by a self-cloning method. Two methods were applied: in one, a stop codon was introduced in the MRU ORF by point mutation; in the other, the entire MRU ORF was deleted from the genome. In both methods, a plasmid vector containing drug-resistance and counter-selectable markers was used to introduce the mutation. We successfully obtained the strains, which did not contain the plasmid sequences, by both methods. Small-scale sake brewing tests using these SC strains (strains obtained by the self-cloning method) found that DMTS-P1 was hardly detected in sake brewed with SC strains, and DMTS production after sake storage was greatly reduced as compared with the parent strain. The components of brewed sake were almost the same between the SC and parent strains. These results suggest that SC strains can produce sake with higher flavor stability without changing the sake brewing properties.