Cryptic Oxidative Transamination of Hydroxynaphthoquinone in Natural Product Biosynthesis

Pyridoxal 5′-phosphate (PLP)-dependent enzymes are a group of versatile enzymes that catalyze various reactions, but only a small number of them react with O₂. Here, we report an unprecedented PLP-dependent enzyme, NphE, that catalyzes both transamination and two-electron oxidation using O₂ as an oxidant. Our intensive analysis reveals that NphE transfers the l-glutamate-derived amine to 1,3,6,8-tetrahydroxynaphthalene-derived mompain to form 8-amino-flaviolin (8-AF) via a highly conjugated quinonoid intermediate that is reactive with O₂. During the NphE reaction, O₂ is reduced to yield H₂O₂. An integrated technique involving NphE structure prediction by AlphaFold v2.0 and molecular dynamics simulation suggested the O₂-accessible cavity. Our in vivo results demonstrated that 8-AF is a genuine biosynthetic intermediate for the 1,3,6,8-tetrahydroxynaphthalene-derived meroterpenoid naphterpin without an amino group, which was supported by site-directed mutagenesis. This study clearly establishes the NphE reaction product 8-AF as a common intermediate with a cryptic amino group for the biosynthesis of terpenoid–polyketide hybrid natural products.