Construction of a synthetic metabolic pathway for biosynthesis of threonine from ethylene glycol.

Ethylene glycol is a promising substrate for bioprocesses which can be derived from widely abundant CO₂ or plastic waste. In this work, we describe the construction of an eight-step synthetic metabolic pathway enabling carbon-conserving biosynthesis of threonine from ethylene glycol. This route extends the previously disclosed synthetic threose-dependent glycolaldehyde assimilation (STEGA) pathway for the synthesis of 2-oxo-4-hydroxybutyrate with three additional reaction steps catalyzed by homoserine transaminase, homoserine kinase, and threonine synthase. We first validated the functionality of the new pathway in an Escherichia coli strain auxotrophic for threonine, which was also employed for discovering a better-performing D-threose dehydrogenase enzyme activity. Subsequently, we transferred the pathway to producer strains and used ¹³C-tracer experiments to improve threonine biosynthesis starting from glycolaldehyde. Finally, extending the pathway for ethylene glycol assimilation resulted in the production of up to 6.5 mM (or 0.8 g L^-1) threonine by optimized E. coli strains at a yield of 0.10 mol mol^-1 (corresponding to 20 % of the theoretical yield).