A gene encoding a DUF523 domain protein is involved in the conversion of 2-thiouracil into uracil

Modified nucleotides are present in many RNA species in all Domains of Life. While the biosynthetic pathways of such nucleotides are well studied, much less is known about the degradation of RNAs and the return to the metabolism of modified nucleotides, their respective nucleosides or heterocyclic bases. Using an E. coli uracil auxotroph, we screened the metagenomic libraries for genes, which would allow the conversion of 2-thiouracil to uracil and thereby lead to the growth on a defined synthetic medium. We show that a gene encoding a protein consisting of previously uncharacterized Domain of Unknown Function 523 (DUF523) is responsible for such phenotype. We have purified this recombinant protein and demonstrated that it contains a FeS cluster. The substitution of cysteines, which have been predicted to form such clusters, with alanines abolished the growth phenotype. We conclude that DUF523 is involved in the conversion of 2-thiouracil into uracil in vivo.