Production of Phenyldiazene Derivatives Using the Biosynthetic Pathway of an Aromatic Diazo Group-Containing Natural Product from an Actinomycete.

The diazo group is an important functional group in organic synthesis because it confers high reactivity to the compounds and has been applied in various chemical reactions, such as the Sandmeyer reaction, Wolff rearrangement, cyclopropanation, and C-N bond formation with active methylene compounds. Previously, we revealed that 3-diazoavenalumic acid (3-DAA), which is potentially produced by several actinomycete species and contains an aromatic diazo group, is a biosynthetic intermediate of avenalumic acid. In this study, we aimed to construct a production system for phenyldiazene derivatives by adding several active methylene compounds to the culture of a 3-DAA-producing recombinant actinomycete. First, acetoacetanilide and its derivatives, which have an active methylene and are raw materials for arylide yellow dyes, were individually added to the culture of a 3-DAA-producing actinomycete. When their metabolites were analyzed, each expected compound with a phenyldiazenyl moiety was detected in the culture extract. Moreover, we established a one-pot in vitro enzymatic production system for the same phenyldiazene derivatives using a highly reactive diazotase, CmaA6. These results showed that the diazo group of natural products is an attractive tool for expanding the structural diversity of natural products both in vivo and in vitro.