Developing bottom-fermenting yeast strains that produce high SO₂ levels by integrated metabolome and transcriptome analysis

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

^* To whom correspondence should be addressed. Email: satoshiy{at}kirin.co.jp.

	Abstract

Sulfite plays an important role in beer flavor stability. Although breeding of bottom-fermenting yeast 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 development of a high 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 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 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 parental levels of H₂S.