Metabolic engineering to eliminate the Crabtree Effect in Saccharomyces cerevisiae: The AlcoFree Yeast.
Arthur L. Kruckeberg
Research & Development, Gothia Yeast Solutions AB, Terrasssgatan 7, Gothenburg, 41133, Sweden
Saccharomyces cerevisiae is a Crabtree-positive yeast: under aerobic conditions it displays fermentative metabolism at high growth rates and extracellular glucose concentrations. For biotechnological applications that require fermentation products (e.g. brewing and baking) this is a desirable trait. However, for production of biomass or biomass components, the Crabtree effect is undesirable, as it results in lower yields and waste of substrate. Crabtree metabolism is conventionally avoided by complex and cumbersome cultivation strategies.Recently, a means of abolishing the Crabtree effect by metabolic engineering has been described (Otterstedt et al, EMBO Reports, in press). This is achieved by replacing the native hexose transporters of yeast with a chimeric transporter that has low transport capacity (the AlcoFree genotype). The glycolytic flux sustained by this transporter is sufficient to maintain respiratory metabolism and a high growth rate, without substantial spillover from pyruvate into ethanolic fermentation. Here the bioprocess characteristics of AlcoFree yeast will be presented under different cultivation regimes: batch, accelerostat, and fed-batch. The results demonstrate that AlcoFree yeast has higher yields of biomass and heterologous protein than wildtype during batch cultivation, and higher critical growth rates and productivity characteristics during accelerostat and fed-batch cultivation. Possible uses of this phenotype in biotechnology are discussed.
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