Metabolomic approach for improving ethanol stress tolerance in Saccharomyces cerevisiae.
The budding yeast Saccharomyces cerevisiae is widely used for brewing and ethanol production. The ethanol sensitivity of yeast cells is still a serious problem during ethanol fermentation, and a variety of genetic approaches (e.g., random mutant screening under selective pressure of ethanol) have been developed to improve ethanol tolerance. In this study, we developed a strategy for improving ethanol tolerance of yeast cells based on metabolomics as a high-resolution quantitative phenotypic analysis. We performed gas chromatography-mass spectrometry analysis to identify and quantify 36 compounds on 14 mutant strains including knockout strains for transcription factor and metabolic enzyme genes. A strong relation between metabolome of these mutants and their ethanol tolerance was observed. Data mining of the metabolomic analysis showed that several compounds (such as trehalose, valine, inositol and proline) contributed highly to ethanol tolerance. Our approach successfully detected well-known ethanol stress related metabolites such as trehalose and proline thus, to further prove our strategy, we focused on valine and inositol as the most promising target metabolites in our study. Our results show that simultaneous deletion of LEU4 and LEU9 (leading to accumulation of valine) or INM1 and INM2 (leading to reduction of inositol) significantly enhanced ethanol tolerance. This study shows the potential of the metabolomic approach to identify target genes for strain improvement of S. cerevisiae with higher ethanol tolerance.
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Erika
E
Ohta
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Yasumune
Y
Nakayama
Department of Bioscience, Nagahama Institute of Bio-Science and Technology, 1266 Tamura-cho, Nagahama, Shiga 526-0829, Japan.
Yukio
Y
Mukai
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Takeshi
T
Bamba
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan. Electronic address: fukusaki@bio.eng.osaka-u.ac.jp.
Eiichiro
E
Fukusaki
D000431:Ethanol / Q000378:metabolism* / Q000494:pharmacology*; D005285:Fermentation / Q000187:drug effects; D008401:Gas Chromatography-Mass Spectrometry; D007294:Inositol / Q000378:metabolism; D055442:Metabolome / Q000235:genetics; D055432:Metabolomics; D009154:Mutation; D010641:Phenotype; D011392:Proline / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D013312:Stress, Physiological / Q000187:drug effects; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014199:Trehalose / Q000378:metabolism; D014633:Valine / Q000378:metabolism
D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D000431:Ethanol; D007294:Inositol; D011392:Proline; D014199:Trehalose; D014633:Valine
Ethanol tolerance; Gas chromatography/mass spectrometry; Metabolomics; Multivariate analysis; Yeast