(474f) Microbial Production of High Value-Added Chiral Hydroxyacids

Hydroxyacids have the potential to serve as useful chiral building block molecules for a diverse range of products, including polyhydroxyalkanoates-based biodegradable plastics and optically-active fine chemicals. Through metabolic engineering efforts, we have constructed unique pathways for the biosynthesis of both enantiomers of 3-hydroxybutyrate (3HB) in E. coli, where the chirality was controlled by utilizing two enzymes of opposite stereoselectivity. We have further profiled several enzyme homologs to promote production, and analyzed 3HB chiralities, in vitro enzyme activities, and intracellular cofactor levels to fully characterize the 3HB pathway. This work led to strains of E. coli that produce enantiopure (R)-3HB and (S)-3HB with titers of both in excess of 2.5 g/L in shake flask cultures within two days. Furthermore, the successfully constructed chiral 3HB pathway provides a feasible production platform for synthesis of other versatile short-chain hydroxyacids. Recent progress has been made on exploiting biosynthetic potential of the 3HB pathway on synthesis of chiral 3-hydroxyvalerate (3HV) with titers of both up to 2.0 g/L as well as 3-hydroxyhexanoate (3HH) and their isomers by either supplying a second carbon source or harnessing the power of natural metabolic pathways. In addition, multiple genetic alternations are being investigated to boost the ratios of higher hydroxyacids to 3HB. This presentation will summarize our engineered microbial system for production of the above-mentioned chiral hydroxyacids.