Simultaneous utilization of glucose and xylose by metabolically engineered Pseudomonas putida for the production of 3-hydroxypropionic acid.

Pseudomonas putida,a robust candidate for lignocellulosicbiomass-based biorefineries, encounters challenges in metabolizing xylose. In this study, Weimberg pathway was introduced intoP. putidaEM42 under a xylose-inducible promoter, resulting in slow cell growth (0.05 h) on xylose.Through adaptive laboratory evolution, an evolved strain exhibited highly enhanced growth on xylose (0.36 h), comparable to that on glucose (0.39 h). Whole genome sequencing identified four mutations, with two key mutations located inPP3380andPP2219. Reverse-engineered strain 8EM42_Xyl, harboring these two mutations, showed enhanced growth on xylose but co-utilizing glucose and xylose at a rate of 0.3 g/L/h. Furthermore, 8EM42_Xyl was employed for 3-hydroxypropionic acid (3HP) production from glucose and xylose by expressing malonyl-CoA reductase and acetyl-CoA carboxylase, yielding 29 g/L in fed-batch fermentation. Moreover, the engineered strain exhibited promising performance in 3HP production from empty palm fruit bunch hydrolysate, demonstrating its potential as a promising cell factory forbiorefineries.