Genomic and Metabolomic Analyses of a Piezosensitive Mutant of Saccharomyces cerevisiae and Application for Generation of Piezosensitive Niigata-Sake Yeast Strains

Abstract

A sparkling-type draft cloudy sake (Japanese rice wine), AWANAMA, was recently developed using high hydrostatic pressure (HHP) treatment as a non-thermal pasteurization method. This prototype sake has a high potential market value, since it retains the fresh taste and flavor similar to draft sake while avoiding over-fermentation. From an economic point of view, a lower pressure level for HHP pasteurization is still required. In this study, we carried out a genome analysis of a pressure-sensitive (piezosensitive) mutant strain, a924E1, which was generated by UV mutagenesis from a laboratory haploid Saccharomyces cerevisiae strain, KA31a. This mutant strain had a deletion of the COX1 gene region in the mitochondrial DNA and had deficient aerobic respiration and mitochondrial functions. A metabolomic analysis revealed restricted flux in the TCA cycle of the strain. The results enabled us to use aerobic respiration deficiency as an indicator for screening a piezosensitive mutant. Thus, we generated piezosensitive mutants from a Niigata-sake yeast strain, S9arg, which produces high levels of ethyl caproate but does not produce urea and is consequently suitable for brewing a high-quality sake. The resultant piezosensitive mutants showed brewing characteristics similar to the S9arg strain. This study provides a screening method for generating a piezosensitive yeast mutant as well as insight on a new way of applying HHP pasteurization.

Importance of Intracellular Energy Status on High-Hydrostatic-Pressure Inactivation of sake Yeast Saccharomyces cerevisiae

“…A laboratory-scale sake brewing test was carried out according to our previous report [ 17 ], which was conducted by Mr. Iwao Takahashi (a master brewer at Kanemasu Sake Co., Shibata, Japan). Yeast strains were pre-cultivated in 3 mL of YPD medium at 30 °C for 24 h with shaking.…”

“…Strain UV1 was cultivated in a YPD medium containing 2% ( w / v ) or 15% ( w / v ) glucose for 48 h at 30 °C, and it was subjected to metabolite extraction. The intracellular metabolites were extracted from cells as described in our previous report [ 17 ]. Briefly, a 5 mL portion of the culture broth was mixed with 7 mL of pre-chilled (−60 °C) 60% ( v / v ) methanol in water.…”

“…Previously, we obtained a pressure-sensitive (piezosensitive) mutant strain cultivated on a conventional YPD medium containing 2% ( w / v ) glucose from S. cerevisiae by random mutagenesis using ultraviolet irradiation (UV) [ 16 ]. Genome analysis revealed that this mutant strain had a deletion of the COX1 gene that encodes subunit I of cytochrome c oxidase in mitochondrial DNA; the strain was deficient in aerobic respiration and/or mitochondrial functions [ 17 ]. We also obtained piezosensitive mutant strains, which also have a deletion in the COX1 gene region in mitochondrial DNA, from a Niigata sake yeast strain, S9arg [ 17 ].…”

“…Genome analysis revealed that this mutant strain had a deletion of the COX1 gene that encodes subunit I of cytochrome c oxidase in mitochondrial DNA; the strain was deficient in aerobic respiration and/or mitochondrial functions [ 17 ]. We also obtained piezosensitive mutant strains, which also have a deletion in the COX1 gene region in mitochondrial DNA, from a Niigata sake yeast strain, S9arg [ 17 ]. These findings suggest that deficiencies in aerobic respiration and/or mitochondrial function cause the piezosensitivity of yeast.…”

Editorial for the Special Issue, “Quality Assay, Processing and Bio-Function of Rice Products”

“…However, further improvements to its brewing process for commercialization still require new HHP techniques at lower pressure levels and the development of pressure-sensitive (piezosensitive) sake yeast strains. Shigematsu et al [ 7 ], in their study ‘The Genomic and Metabolomic Analyses of a Piezosensitive Mutant of Saccharomyces cerevisiae and Application for Generation of Piezosensitive Niigata-Sake Yeast Strains’, provide a screening method for generating a piezosensitive yeast mutant as well as insight into a new method of applying HHP pasteurization. Food quality and safety are important drivers in consumers preferences, and they also govern rice production and processing.…”