Towards the conversion of CO(2)into optically pureN-carbamoyl-l-aspartate and orotate by anin vitromulti-enzyme cascade

摘要

A novelin vitromulti-enzymatic system was constructed for the conversion of CO(2)into optically pureN-carbamoyl-l-aspartate and orotate. Four cascade enzymes, carbamoyl phosphate synthase (CPS), aspartate carbamoyl-transferase (ATCase), dihydroorotase (DHOase), and dihydroorotate dehydrogenase (DHOD), were used for the biotransformation of feedstocks into products in one pot while polyphosphate kinase (PPK) was coupled for ATP regeneration. Under the optimal reaction conditions and substrate concentrations, 19.2 mMN-carbamoyl-l-aspartate was produced within 3.0 h, which enabled the synthesis of 15.5 mM orotate. The yields ofN-carbamoyl-l-aspartate and orotate based on the addedl-aspartic acid reached 96.0% and 93.3%, respectively. This work highlights the ease of producing optically pureN-carbamoyl-l-aspartate and orotateviaanin vitrobiomanufacturing platform and provides a novel pathway for the utilization of renewable feedstock for the synthesis of value-added commodity chemicals.