Controlled Expression of the Dominant Flocculation Genes FLO1 , FLO5 , and FLO11 in Saccharomyces cerevisiae

Abstract

In many industrial fermentation processes, the Saccharomyces cerevisiae yeast should ideally meet two partially conflicting demands. During fermentation, a high suspended yeast count is required to maintain a satisfactory rate of fermentation, while at completion, efficient settling is desired to enhance product clarification and recovery. In most fermentation industries, currently used starter cultures do not satisfy this ideal, probably because nonflocculent yeast strains were selected to avoid fermentation problems. In this paper, we assess molecular strategies to optimize the flocculation behavior of S. cerevisiae. For this purpose, the chromosomal copies of three dominant flocculation genes, FLO1, FLO5, and FLO11, of the haploid nonflocculent, noninvasive, and non-flor-forming S. cerevisiae FY23 strain were placed under the transcriptional control of the promoters of the ADH2 and HSP30 genes. All six promoter-gene combinations resulted in specific flocculation behaviors in terms of timing and intensity. The strategy resulted in stable expression patterns providing a platform for the direct comparison and assessment of the specific impact of the expression of individual dominant FLO genes with regard to cell wall characteristics, such as hydrophobicity, biofilm formation, and substrate adhesion properties. The data also clearly demonstrate that the flocculation behavior of yeast strains can be tightly controlled and fine-tuned to satisfy specific industrial requirements.

125th Anniversary Review: Yeast Flocculation and Sedimentation in Brewing

“…FLO5-based constructs were less efficient but still caused significant flocculation. FLO11-based constructs caused only weak flocculation, but instead caused strong flor formation and invasive growth, which were not observed with FLO1 and FLO5 constructs 20 .…”

“…In contrast, several studies 10,16,20,102,109 show that the timing and strength of flocculation can be successfully controlled in a planned way by genetic engineering, although it may sometimes be difficult to find the tools (promoters etc.) giving the required behaviour (see, e.g.…”

“…In the work of Govender et al 20 , FLO1, FLO5 and FLO11 were put under the control of ADH2 or HSP30 promoters by replacing the native promoters of these genes in a haploid, non-flocculent laboratory strain. ADH2 or HSP30 promoters are strongly induced when cells enter the stationary phase.…”

Flocculation in ale brewing strains of Saccharomyces cerevisiae: re-evaluation of the role of cell surface charge and hydrophobicity

“…An increase of yeast surface hydrophobicity when the glycoproteins Flo1p, Flo5p, Flo9p and Flo10p are present in yeast cell walls (Verstrepen et al 2001;Govender et al 2008;Mulders et al 2009;Govender et al 2010) was described. Similarly, the expression of FLO11 also confers hydrophobicity to yeast cells (Guo et al 2000;Reynolds and Fink 2001;Fidalgo et al 2006;Mortensen et al 2007;Govender et al 2008;Mulders et al 2009).…”

“…Similarly, the expression of FLO11 also confers hydrophobicity to yeast cells (Guo et al 2000;Reynolds and Fink 2001;Fidalgo et al 2006;Mortensen et al 2007;Govender et al 2008;Mulders et al 2009). Nevertheless, contradictory results regarding the flocculation behaviour of FLO11 transformants have been presented.…”