Evidence that J-binding protein 2 is a thymidine hydroxylase catalyzing the first step in the biosynthesis of DNA base J

The genomic DNA of kinetoplastid parasites contains a unique modified base. (beta-D-glucosylhydroxymethyluracil or base J. We recently reported that two proteins, called J-binding protein (JBP) 1 and 2, which regulate the levels of J in the genome, display features of the family of Fe(II)-2-oxoglutarate dependent dioxygenases and are likely to be the enzymes catalyzing the first step in J biosynthesis. In this study, we examine the effects of replacing the four conserved residues critical for the activity of this class of enzymes on the function of Leishmania tarentolae JBP2. The results show that each of these four residues is indispensable for the ability of JBP2 to stimulate J synthesis, while mutating non-conserved residues has no consequences. We conclude that JBP2, like JBP1, is in all probability a thymidine hydroxylase involved in the biosynthesis of base J. (C) 2008 Elsevier B.V. All rights reserved.