Development of a sustainable approach to produce galactaric acid from sunflower heads by engineered Pseudomonas putida KT2440

Upgrading of agricultural inferior resources to produce bio-based chemicals through biotechnological approaches has gained considerable attention in recent years. Galactaric acid (GA), serving as an intriguing building block for a series of Polymers, can be synthesized from d-galacturonic acid (GalA) stemmed from agricultural waste, but the biological synthesis approach should be further improved to compete with Other methods. In this work, GA biosynthesis depending on the intrinsic pathway of Pseudomonas putida KT2440 was investigated for the first time, and a sustainable approach was developed to produce GA from GalA derived from sunflower head pectin by engineered P. putida. Firstly, crude GalA was obtained by enzymatic hydrolysis of sunflower head pectin. Subsequently, engineered P. putida strains blocking GA degradation route were constructed based on the identification and inactivation of the related metabolic genes of GA. Furthermore, a uronate dehydrogenase (UDH) and a pyrroloquinoline quinone-dependent glucose dehydrogenase responsible for the oxidization of GalA were identified. Finally, P. putida ΔPP_2834 ΔPP_3601 udh, which deactivated the GA degradation activity and heterologously expressed the UDH, was observed to produce GA at a titer of 22.40 g/L and 7.36 g/L from commercial GalA (20 g/L) and crude GalA (10 g/L), respectively, which demonstrated the feasibility and practicality of this green approach.

Biosynthesis; Galactaric acid; Pseudomonas putida; Sunflower heads; d-Galacturonic acid.