Genetically Engineering Escherichia coli to Produce Xylitol from Corncob Hydrolysate without Lime Detoxification

Abstract

Before fermentation with hemicellulosic hydrolysate as a substrate, it is generally necessary to detoxify the toxic substances that are harmful to microorganism growth. Cyclic AMP receptor protein (CRP) is a global regulator, and mutation of its key sites may have an important impact on E. coli virulence tolerance. Using corncob hydrolysate without ion-exchange or lime detoxification as the substrate, shake flask fermentation experiments showed that CRP mutant IS5-dG (I112L, T127G, A144T) produced 18.4 g/L of xylitol within 34 h, and the OD600 was 9.7 at 24 h; these values were 41.5% and 21.3% higher than those of the starting strain, IS5-d, respectively. This mutant produced 82 g/L of xylitol from corncob hydrolysate without ion-exchange or lime detoxification during fed-batch fermentation in a 15-L bioreactor, with a productivity of 1.04 g/L/h; these values were 173% and 174% higher than the starting strain, respectively. To our knowledge, this is the highest xylitol concentration and productivity produced by microbial fermentation using completely non-detoxified hemicellulosic hydrolysate as the substrate to date. This study also showed that alkali neutralization, high temperature sterilization, and fermentation of the hydrolysate had important effects on the xylose loss rate and xylitol production.

Valorization of Corn Cobs for Xylitol and Bioethanol Production through Column Reactor Process

“…Specifically, the genetically modified Saccharomyces cerevisiae PE-2-GRE3 demonstrated a high productivity of 0.83 g/L•h [33]. Similarly, engineered Escherichia coli achieved an impressive productivity of up to 1.04 g/L•h, setting a record for microbial fermentation [34].…”

“…In fact, research on the production of xylitol from corncobs primarily relies on the use of genetically modified microorganisms (GMOs) engineered to utilize xylose as a carbon source [32][33][34]. Specifically, the genetically modified Saccharomyces cerevisiae PE-2-GRE3 demonstrated a high productivity of 0.83 g/L•h [33].…”

Metabolic engineering in lignocellulose biorefining for high-value chemicals: recent advances, challenges, and outlook for enabling a bioeconomy

“…In a subsequent study, they mutated the CRP gene in the above E. coli strain to improve the inhibitory tolerance. The newly engineered E. coli mutant (I112L, T127G, A144T) could produce 82 g/L xylitol from un-detoxified condensed corncob hydrolysate via fed-batch fermentation, whereas host-engineered E. coli only produced 30 g/L xylitol (Yuan et al, 2023). Similarly, another mutant (I112L, T127I, and A144T) produced 164 g/L xylitol from detoxified corncob hydrolysate (Yuan et al, 2020).…”