Modifications of diketopiperazines assembled by cyclodipeptide synthases with cytochrome P₄₅₀ enzymes

2,5-Diketopiperazines are the smallest cyclic peptides comprising two amino acids connected via two peptide bonds. They can be biosynthesized in nature by two different enzyme families, either by nonribosomal peptide synthetases or by cyclodipeptide synthases. Due to the stable scaffold of the diketopiperazine ring, they can serve as precursors for further modifications by different tailoring enzymes, such as methyltransferases, prenyltransferases, oxidoreductases like cyclodipeptide oxidases, 2-oxoglutarate-dependent monooxygenases and cytochrome P₄₅₀ enzymes, leading to the formation of intriguing secondary metabolites. Among them, cyclodipeptide synthase-associated P₄₅₀s attracted recently significant attention, since they are able to catalyse a broader variety of astonishing reactions than just oxidation by insertion of an oxygen. The P₄₅₀-catalysed reactions include hydroxylation at a tertiary carbon, aromatisation of the diketopiperazine ring, intramolecular and intermolecular carbon-carbon and carbon-nitrogen bond formation of cyclodipeptides and nucleobase transfer reactions. Elucidation of the crystal structures of three P₄₅₀s as cyclodipeptide dimerases provides a structural basis for understanding the reaction mechanism and generating new enzymes by protein engineering. This review summarises recent publications on cyclodipeptide modifications by P₄₅₀s.Key Points• Intriguing reactions catalysed by cyclodipeptide synthase-associated cytochrome P₄₅₀s• Homo- and heterodimerisation of diketopiperazines• Coupling of guanine and hypoxanthine with diketopiperazines.