Control of the galactose-to-glucose consumption ratio in co-fermentation using engineered Escherichia coli strains

Abstract

Marine biomasses capable of fixing carbon dioxide have attracted attention as an alternative to fossil resources for fuel and chemical production. Although a simple co-fermentation of fermentable sugars, such as glucose and galactose, has been reported from marine biomass, no previous report has discussed the fine-control of the galactose-to-glucose consumption ratio in this context. Here, we sought to finely control the galactose-to-glucose consumption ratio in the co-fermentation of these sugars using engineered Escherichia coli strains. Toward this end, we constructed E. coli strains GR2, GR2P, and GR2PZ by knocking out galRS, galRS-pfkA, and galRS-pfkA-zwf , respectively, in parent strain W3110. We found that strains W3110, GR2, GR2P, and GR2PZ achieved 0.03, 0.09, 0.12, and 0.17 galactose-to-glucose consumption ratio (specific galactose consumption rate per specific glucose consumption rate), respectively, during co-fermentation. The ratio was further extended to 0.67 by integration of a brief process optimization for initial sugar ratio using GR2P strain. The strategy reported in this study will be helpful to expand our knowledge on the galactose utilization under glucose conditions.

Effect of deregulation of repressor-specific carbon catabolite repression on carbon source consumption in Escherichia coli

“…In a recent report, it was reported that the wild-type E. coli W3110 cells first consumed glucose rapidly with a rate of 1.37 g/gDCW/h, followed by galactose consumption with a rate of 0.04 g/gDCW/h in both sugars co-fermentation [20]. This indicates that galactose consumption was strictly inhibited by the CCR mechanism in the W3110 strain (Fig.…”

“…1). In the previous report, under the glucose and galactose co-fermentation (each 4 g/L), the E. coli GR2 cells showed a similar specific glucose consumption rate of the W3110 strain [20]. However, in the same co-fermentation, the GR2 strain achieved a 3.38 times increase in specific galactose consumption rate [20].…”

“…In this study, the wild-type E. coli W3110 and its mutants GR2, GR2P, and GR2PZ strains were used in order to see the effect of deregulation of repressor-specific CCR. The mutants GR2 (galR − and galS − ), GR2P (galR − , galS − , and pfkA − ), and GR2PZ (galR − , galS − , pfkA − , and zwf − ) were constructed by knockout of the genes using Lambda Red and Cre recombinases in our previous study [20].…”

“…Galactose is assumed as a valuable carbon source found in marine macroalgae for bio-based chemical production [19]. For this reason, our recent study has reported a strategy to control two carbon source consumption in the glucose and galactose co-fermentation by altering the CCR mechanism in E. coli [20]. In the previous report, mutant E. coli strains GR2 (galR − , galS − ), GR2P (galR − , galS − , pfkA − ), and GR2PZ (galR − , galS − , pfkA − , zwf − ) were intensively characterized focusing on the co-fermentation of glucose and galactose [20].…”

“…For this reason, our recent study has reported a strategy to control two carbon source consumption in the glucose and galactose co-fermentation by altering the CCR mechanism in E. coli [20]. In the previous report, mutant E. coli strains GR2 (galR − , galS − ), GR2P (galR − , galS − , pfkA − ), and GR2PZ (galR − , galS − , pfkA − , zwf − ) were intensively characterized focusing on the co-fermentation of glucose and galactose [20]. However, these mutant E. coli strains have not yet been characterized in fermentations supplemented with sole carbon source.…”