Identification and mechanistic analysis of a bifunctional enzyme involved in the C-N and N-N bond formation

N-N bonded compounds are a relatively small but valuable group of natural products with a variety of important biological activities, including anti-inflammatory, anti-tumor, anti-bacterial and anti-oxidant activities. Currently, the synthesis of natural products containing N-N bonds is mainly based on chemical synthesis, but the chemical synthesis of N-N bonds may have safety issues and raw material sustainability problems. A variety of N-N bond biosynthetic mechanisms exist in nature, including comproportionation, rearrangement, and radical recombination reactions, which provide ideas for the biosynthesis of many N-N bond compounds. But at this stage, only a few N-N bond synthases have been reported, and difficulties exist in the biosynthesis of N-N bond-containing compounds. The new N-N-bond synthase PAI2, which was found to catalyze the synthesis of piperazinic acid, expands the family of N-N-bond synthases and provides better catalytic activity and different enzymatic properties for the substrate L-N5-hydroxyornithine than the known enzyme KtzT derived from the piperazinic acid biosynthetic gene cluster of Kutzneria sp. 744. Moreover, PAI2 was found to have the ability to catalyze the formation of C-N bonds in Pro.