Presentation Information
[P02-183]Comprehensive analysis of rice-koji proteins from Aspergillus oryzae and their effects on the sake metabolome
○Ryousuke Kataoka1, Shinichiro Fukuhara1,2, Shingo Kakiuchi1,2, Minori Kono1, Sharon Marie Bahena-Garrido1, Yuko Komatsu-Hata1, Kazuhiro Iwashita1,2,3 (1. National Research Institute of Brewing (Japan), 2. Department of Molecular Biotechnology, Graduate School of Advanced Science of Matter, Hiroshima University (Japan), 3. Unit of Biotechnology, Graduate School of Integrated Sciences for Life, Hiroshima University (Japan))
Keywords:
Aspergillus oryzae,Japanese sake,metabolome,rice-koji
[Background]
Sake is a traditional Japanese alcoholic beverage brewed from rice, rice-koji, and water. Its production relies on the coordinated activities of Aspergillus oryzae and Saccharomyces cerevisiae in a process known as multiple parallel fermentation, in which rice starch is saccharified by A. oryzae and the resulting sugars are simultaneously fermented into alcohol by yeast. Because the flavor and chemical composition of sake are shaped by a complex interplay among raw materials, microorganisms, and fermentation conditions, understanding the contributions of microbial factors is essential for improving brewing technology. Rice-koji, produced by growing A. oryzae on steamed rice, is therefore a key component of sake brewing. In addition to well-known starch-degrading and proteolytic enzymes, A. oryzae is thought to produce many other proteins during rice-koji fermentation, but the full range of these proteins and their specific roles in sake brewing remain largely unclear.
[Purpose and Results]
In this study, we sought to comprehensively identify proteins expressed by A. oryzae in rice-koji and to evaluate their effects on sake components. Proteomic analyses of four types of rice-koji identified 159 rice-koji proteins (RKPs), corresponding to 159 rkp genes. Based on these candidates, selected genes were disrupted individually, resulting in 85 distinct Δrkp strains. Koji-making experiments revealed alterations in the mycelial biomass of A. oryzae in rice-koji, and small-scale sake brewing experiments using these disruptants were associated with changes in multiple sake components. Furthermore, sake metabolomic analysis showed that disruption of 57 rkp genes caused marked changes in at least one metabolite peak detected in sake.
[Conclusion]
These results demonstrate that proteins expressed by A. oryzae during rice-koji production have broad effects on fungal physiology and the sake metabolome. This study provides a framework for understanding how rice-koji proteins contribute to sake brewing and for prioritizing candidate factors for further functional analysis.
Sake is a traditional Japanese alcoholic beverage brewed from rice, rice-koji, and water. Its production relies on the coordinated activities of Aspergillus oryzae and Saccharomyces cerevisiae in a process known as multiple parallel fermentation, in which rice starch is saccharified by A. oryzae and the resulting sugars are simultaneously fermented into alcohol by yeast. Because the flavor and chemical composition of sake are shaped by a complex interplay among raw materials, microorganisms, and fermentation conditions, understanding the contributions of microbial factors is essential for improving brewing technology. Rice-koji, produced by growing A. oryzae on steamed rice, is therefore a key component of sake brewing. In addition to well-known starch-degrading and proteolytic enzymes, A. oryzae is thought to produce many other proteins during rice-koji fermentation, but the full range of these proteins and their specific roles in sake brewing remain largely unclear.
[Purpose and Results]
In this study, we sought to comprehensively identify proteins expressed by A. oryzae in rice-koji and to evaluate their effects on sake components. Proteomic analyses of four types of rice-koji identified 159 rice-koji proteins (RKPs), corresponding to 159 rkp genes. Based on these candidates, selected genes were disrupted individually, resulting in 85 distinct Δrkp strains. Koji-making experiments revealed alterations in the mycelial biomass of A. oryzae in rice-koji, and small-scale sake brewing experiments using these disruptants were associated with changes in multiple sake components. Furthermore, sake metabolomic analysis showed that disruption of 57 rkp genes caused marked changes in at least one metabolite peak detected in sake.
[Conclusion]
These results demonstrate that proteins expressed by A. oryzae during rice-koji production have broad effects on fungal physiology and the sake metabolome. This study provides a framework for understanding how rice-koji proteins contribute to sake brewing and for prioritizing candidate factors for further functional analysis.
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