Integration of metabolic and bioprocess engineering for the production of β-ketoadipic acid from glucose and xylose by Pseudomonas putida

β-Ketoadipic acid is a common intermediate in aerobic microbial aromatic catabolism that can be used as a monomer in performance-advantaged biopolymers. Here, we engineered Pseudomonas putida KT2440 to produce β-ketoadipate from glucose and xylose, the most prevalent carbohydrates derived from lignocellulosic polysaccharides. With the top-performing strain, P. putida GR038, we conducted bioprocess development using glucose and xylose as substrates in a 2 : 1 molar ratio to mimic lignocellulosic hydrolysate. Fed-batch cultivations achieved a titer of 65.8 g L⁻¹ and a rate of 0.69 g L⁻¹ h⁻¹, with a C-mol yield of 0.52. Application of adsorptive in situ product recovery further improved the effective titer to 92.0 g L⁻¹ and the rate to 0.83 g L⁻¹ h⁻¹, while also improving the downstream purity of β-ketoadipate from 88.3 wt% to 99.0 wt%. These results show promise towards industrial production of β-ketoadipate from lignocellulosic sugars.