Metabolic Engineering 11

Modulation of Carbon Preference in Enterobacter Aerogenes for Efficient Utilization of Sugarcane Molasses for 2,3-Butanediol Production

Author

Oh, M. K., Korea university

Due to its cost-effectiveness and rich sugar composition, sugarcane molasses is considered to be a promising carbon source for biorefinery. However, the sugar mixture in sugarcane molasses is not consumed as efficiently as glucose in microbial fermentation due to complex interactions among their utilizing pathways, such as carbon catabolite repression (CCR). Therefore, E. aerogenes was engineered to alleviate CCR and improve sugar utilization by modulating its carbon preference. To improve sucrose utilization in E. aerogenes, a sucrose regulator (ScrR) was deleted because sucrose occupies more than half of the sugar in the molasses. The deletion mutation increased the sucrose consumption rate significantly when sucrose or sugarcane molasses was used as a carbon source, but the efficiency of fructose utilization in the scrR mutant was relatively reduced. Therefore, the gene encoding catabolite repressor/activator (Cra) was removed to increase the fructose consumption rate. The disruption of cra improved the consumption rate of fructose, while significantly decreasing sucrose utilization, resulting in the accumulation of sucrose in the fermentation medium. Cra regulation on expression of the scrAB operon involved in sucrose catabolism was verified by reverse transcription and real-time PCR, and the efficiency of sucrose utilization was restored by disrupting the scrR gene and overexpressing the scrAB operon. In addition, the overexpression of ptsG gene involved in glucose utilization enhanced the glucose preference among mixed sugars, which relieved glucose accumulation in fed-batch fermentation. These genetic modifications enabled the mutant strain to metabolize all sugars in sugarcane molasses efficiently and simultaneously, which increased fermentation duration and relieved the accumulation of certain carbon sources in fed-batch fermentation, and the maximum titer of 2,3-butanediol production by the mutant reached 140.0 g/L at 54 h.