(15e) Establishing a Platform Escherichia coli Strain to Generate Xylose-Derived Value-Added Products

Wang, J., University of Georgia
Shen, X., University of Georgia
Yuan, Q., Beijing University of Chemical Technology
Yan, Y., University of Georgia
Xylose is the most abundant C5 sugar in lignocellulosic biomass and also represents a source of carbon from non-edible feedstocks. Here, we report the construction of a platform Escherichia coli strain for the production of high value compounds via the nonphosphorylative xylose metabolism. Firstly, a 3,4-dihydroxybutanal over-producing pathway was constructed by efficient enzymes screening and host strain engineering. Then two 3,4-dihydroxybutanal dehydrogenases were identified to efficiently convert 3,4-dihydroxybutanal into 3,4-dihydroxybutyric acid (3,4-DHBA), which is the hydrolyzed form of 3-Hydroxy-γ-butyrolactone (3HBL). 3HBL is one of the top value-added building block for synthesis of various drugs and nutraceuticals. This novel 3,4-DHBA biosynthetic pathway produced 1.27 g/L of 3,4-DHBA in shake flasks, which is the highest titer reported so far. The application of this platform was further demonstrated by building an artificial pathway for biosynthesis of 1,4-butanediol (1,4-BDO). The 3,4-dihydroxybutanal over-producing strain was transformed into a 1,2,4-butanetriol (1,2,4-BTO) over-producing strain by over-expression of endogenous alcohol dehydrogenase, which enabled 1.5 g/L 1,2,4-BTO produced from xylose. Then the Klebsiella oxytoca diol dehydratase was engineered to achieve non-native catalysis of 1,2,4-BTO into 1,4-BDO by a series of rational protein engineering strategies. Those efforts enabled 209 mg/L 1,4-BDO produced via a novel metabolic route from xylose. This work demonstrates great potential for large-scale production of 3,4-DHBA and sets an example to build novel biosynthetic pathways via rational protein engineering.