Use of Genetic Transformation for the Introduction of Flocculence into Yeast

Barney, M. C.; Jansen, Gregor; Helbert, J.

doi:10.1094/asbcj-38-0071

Cited by 10 publications

(9 citation statements)

References 19 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In their seminal study, Barney et al 34) introduced the FLO1 gene into spheroplasted S. cerevisiae to confer the flocculation phenotype. The protoplast fusion technique has also been used in the construction of hybrids between nonflocculent and flocculent strains.…”

Section: Discussionmentioning

confidence: 99%

Construction of FlocculentKluyveromyces marxianusStrains Suitable for High-Temperature Ethanol Fermentation

Nonklang

Ano²,

Abdel-Banat

et al. 2009

Bioscience, Biotechnology, and Biochemistry

View full text Add to dashboard Cite

Flocculating yeasts are highly useful in fermentation processes because these cells can be separated easily from the fermentation mash. However, native yeasts are usually non-flocculating, including Kluyveromyces marxianus, which exhibits a potent high-temperature ethanol fermentation ability. We describe here the construction of flocculent K. marxianus strains via the introduction of the FLO1, FLO5, FLO9, and FLO10 genes from Saccharomyces cerevisiae. The S. cerevisiae FLO genes were overexpressed by upstream insertion of the constitutive TDH3 promoter, resulting in flocculent S. cerevisiae strains. These TDH3p-FLO sequences were then amplified by PCR and introduced directly into a K. marxianus strain. These K. marxianus strains showed a flocculation phenotype, indicating that the introduced S. cerevisiae TDH3 promoter and all FLO genes were functional in this strain. Moreover, a flocculating K. marxianus strain showed the same ethanol production profile as that of its wild-type parent. The K. marxianus flocculating strains we generated should be useful in the future development of cost-effective fed-batch and continuous fermentation systems at high temperatures.Key words: Kluyveromyces marxianus; yeast; flocculation; ethanol fermentationFlocculation is an attractive property in industrial yeasts because the cells can be easily separated from a fermentation mash. The application of flocculating strains to industrial fermentation processes removes the need for extensive and costly cell separation equipment, and these yeasts are thus suitable for both fedbatch and continuous fermentation. [1][2][3][4] In addition, flocculating yeast cells are useful in immobilized cell systems with no inert support materials. 5) However, although flocculation mechanisms and the introduction of this property to non-flocculent yeast strains have been extensively studied in S. cerevisiae, few studies of this nature have been undertaken in other yeast species. 6,7) The mechanism of yeast flocculation involves the interaction of lectin-like proteins, called flocculins, with receptors on a neighboring cell wall, resulting in the formation of aggregates. [8][9][10][11] Environmental factors such as pH, calcium, and organic stress also affect flocculation. 9,[12][13][14] The five FLO genes, FLO1, FLO5, FLO9, FLO10, and FLO11, have been described in S. cerevisiae thus far. [15][16][17][18][19] The FLO1, 5, 9, and 10 genes confer cell-cell adhesion (flocculation) ability, whereas FLO11 is responsible for substrate adhesion. 20) FLO1, FLO5, and FLO9 are highly similar to each other, but FLO10 has lower similarity. The mechanisms of flocculation in other yeasts appear to be lectin-based but of differing specificities. For instance, galactose and its derivatives have been found to inhibit flocculation of K. bulgaricus and K. lactis, suggesting that this interaction involves a galactose-specific lectin. 21,22) The advantages of using flocculating yeast cells in fermentation processes have led to many studies on the construction of flocculent s...

show abstract

Section: Discussionmentioning

confidence: 99%

Construction of FlocculentKluyveromyces marxianusStrains Suitable for High-Temperature Ethanol Fermentation

Nonklang

Ano²,

Abdel-Banat

et al. 2009

Bioscience, Biotechnology, and Biochemistry

View full text Add to dashboard Cite

show abstract

“…The FLO1 gene was constitutively expressed, thereby creating transgenic yeast strains that exhibited a constitutive flocculation property irrespective of the growth phase (2,23,66,67). However, efficient fermentation requires a high suspended cell count, and constitutively flocculating yeast may lead to sluggish or stuck fermentations.…”

Section: Vol 74 2008 Controlled Expression Of Dominant Flocculationmentioning

confidence: 99%

“…In previous attempts to modify flocculation behavior, the flocculation genes FLO1 and FLO5 were introduced into nonflocculent S. cerevisiae brewing yeast strains (2,23,(65)(66)(67). However, the resultant modified yeast strains flocculated constitutively and displayed reduced fermentation performance or increased fermentation times.…”

mentioning

confidence: 99%

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

Gasqui

Domingo

Bester

et al. 2008

Appl Environ Microbiol

120

View full text Add to dashboard Cite

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.

show abstract

“…In 1980, Barney and collaborators [101] Aiming at improving the yield of the batch process in cane molasses fermentation and using a flocculent yeast for a continuous molasses or cane juice fermentation, Figueroa et al [102] reported the transfer of the flocculation property to the yeast S. cerevisiae. This yeast is used as distillery yeast in batch processes in Argentina for ethanol production from sugar cane molasses.…”

Section: Transfer Of Flocculation Properties To Nonflocculent Yeastsmentioning

confidence: 99%

Applications of yeast flocculation in biotechnological processes

Domingues

Vicente

Lima

et al. 2000

Biotechnol. Bioprocess Eng.

View full text Add to dashboard Cite

A review on the main aspects associated with yeast flocculation and its application in biotechnological processes is presented. This subject is addressed following three main aspects -the basics of yeast flocculation, the development of "new" flocculating yeast strains and bioreactor development. In what concerns the basics of yeast flocculation, the state of the art on the most relevant aspects of mechanism, physiology and genetics of yeast flocculation is reported. The construction of flocculating yeast strains includes not only the recombinant constitutive flocculent brewer's yeast, but also recombinant flocculent yeast for lactose metabolisation and ethanol production. Furthermore, recent work on the heterologous β-galactosidase production using a recombinant flocculent Saccharomyces cerevisiae is considered. As bioreactors using flocculating yeast cells have particular properties, mainly associated with a high solid phase hold-up, a section dedicated to its operation is presented. Aspects such as bioreactor productivity and culture stability as well as bioreactor hydrodynamics and mass transfer properties of flocculating cell cultures are considered. Finally, the paper concludes describing some of the applications of high cell density flocculation bioreactors and discussing potential new uses of these systems.

show abstract

Use of Genetic Transformation for the Introduction of Flocculence into Yeast

Cited by 10 publications

References 19 publications

Construction of FlocculentKluyveromyces marxianusStrains Suitable for High-Temperature Ethanol Fermentation

Construction of FlocculentKluyveromyces marxianusStrains Suitable for High-Temperature Ethanol Fermentation

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

Applications of yeast flocculation in biotechnological processes

Contact Info

Product

Resources

About