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Flavin-dependent halogenases(FDHs)are well known to introduce carbon halide bonds(mainly C-Cl and C-Br)into natural products with the assistance of a partner protein flavin reductase to generate reduced flavin(FADH2 or FMNH2).Compared with the common chloride-and bromide-containing natural products(approximately 5,000 compounds),iodinated natural products(approximately 100 compounds)are very limited.Specific iodinases have also rarely been identified in nature to date.This study discovered a novel relationship between iodination and flavin reductases for the first time.Through mechanistic studies,it was identified that peroxide(H2O2)was released from the uncoupling reaction of flavin reductases and then reacted with iodide ions(I)to produce hypoiodous acid(IOH)for the final iodination.Furthermore,this study also unintentionally verified that the recently reported flavin-dependent iodinase VirXl from the marine virus and its two homologs(MBG and NCV)did not catalyze iodination in the in vitro biochemical system but likely belonged to a new phylogenetic clade in the tryptophan halogenase superfamily.As a consequence,actual flavin-dependent iodinases in nature remain to be discovered by the scientific community in the future.