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Applied and Environmental Microbiology, May 2008, p. 2787-2796, Vol. 74, No. 9
0099-2240/08/$08.00+0     doi:10.1128/AEM.01781-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Development of Bottom-Fermenting Saccharomyces Strains That Produce High SO2 Levels, Using Integrated Metabolome and Transcriptome Analysis{triangledown}

Satoshi Yoshida,1* Jun Imoto,2 Toshiko Minato,1 Rie Oouchi,1 Mao Sugihara,3 Takeo Imai,3 Tatsuji Ishiguro,1 Satoru Mizutani,1 Masaru Tomita,2 Tomoyoshi Soga,2 and Hiroyuki Yoshimoto1

Central Laboratories for Frontier Technology, KIRIN Holdings Co., Ltd., 1-13-5 Fukuura Kanazawa-ku, Yokohama-shi, Kanagawa 236-0004, Japan,1 Institute for Advanced Biosciences, Keio University, 246-2 Mizukami Kakuganji, Tsuruoka-shi, Yamagata 997-0052, Japan,2 Research Laboratories for Brewing, KIRIN Brewery Co., Ltd., 1-17-1 Namamugi Tsurumi-ku, Yokohama-shi, Kanagawa 230-8628, Japan3

Received 1 August 2007/ Accepted 24 February 2008

Sulfite plays an important role in beer flavor stability. Although breeding of bottom-fermenting Saccharomyces strains that produce high levels of SO2 is desirable, it is complicated by the fact that undesirable H2S is produced as an intermediate in the same pathway. Here, we report the development of a high-level SO2-producing bottom-fermenting yeast strain by integrated metabolome and transcriptome analysis. This analysis revealed that O-acetylhomoserine (OAH) is the rate-limiting factor for the production of SO2 and H2S. Appropriate genetic modifications were then introduced into a prototype strain to increase metabolic fluxes from aspartate to OAH and from sulfate to SO2, resulting in high SO2 and low H2S production. Spontaneous mutants of an industrial strain that were resistant to both methionine and threonine analogs were then analyzed for similar metabolic fluxes. One promising mutant produced much higher levels of SO2 than the parent but produced parental levels of H2S.

* Corresponding author. Mailing address: Central Laboratories for Frontier Technology, KIRIN Holdings Co., Ltd., 1-13-5 Fukuura Kanazawa-ku, Yokohama-shi, Kanagawa 236-0004, Japan. Phone: 81-45-330-9003. Fax: 81-45-788-4042. E-mail: satoshiy{at}kirin.co.jp

{triangledown} Published ahead of print on 29 February 2008.

Applied and Environmental Microbiology, May 2008, p. 2787-2796, Vol. 74, No. 9
0099-2240/08/$08.00+0     doi:10.1128/AEM.01781-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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