Discovery of a novel gene, involved in the conversion of 2-thiouracil into uracil

Abstract

Modified nucleotides are present in many RNA species in all Domains of Life. The biosynthetic pathways of such nucleotides are well studied. However, much less is known about the degradation of RNAs and the salvage of modified nucleotides, their respective nucleosides or heterocyclic bases to the metabolism. Using an E. coli uracil auxotrophic strain, 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 novel gene encoding previously uncharacterized Domain of Unknown Function (DUF) 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 bind such clusters, with alanines abolished the growth phenotype. We conclude that this domain is required for conversion of 2-thiouracil into uracil in vivo. This work is supported by the Research Council of Lithuania (LMT, MIP-103/2015).