Enzymatic Synthesis of Kynurenic Acid-Linked Dipeptides with C-Terminal Amide

Kynurenic acid (KYNA) ester/amide derivatives were explored for improved biological potency. Herein, we report the discovery of a nonribosomal peptide synthase and P450 enzyme cascade capable of synthesizing KYNA-linked dipeptides in Aspergillus nidulans. In vitro assay together with isotope feeding experiments reveals an intriguing rearrangement from diketopiperazine (DKP) to dipeptide with a C-terminal amide. Feeding tryptophan derivatives allows the generation of various KYNA ester/amide derivatives. Our findings reveal the natural enzymatic machinery for KYNA amide biosynthesis, establishing a foundation for enzymatic access to KYNA amide derivatives.