The domain architecture of the protozoan protein J-DNA-binding protein 1 suggests synergy between base J DNA binding and thymidine hydroxylase activity.

Athanassios Adamopoulos, Tatjana Heidebrecht, Jeroen Roosendaal, Wouter G Touw, Isabelle Q Phan, Jos Beijnen, Anastassis Perrakis

The Journal of biological chemistry 2019 Aug 23

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J-DNA-binding protein 1 (JBP1) contributes to the biosynthesis and maintenance of base J (β-d-glucosyl-hydroxymethyluracil), an epigenetic modification of thymidine (T) confined to pathogenic protozoa such as Trypanosoma and Leishmania JBP1 has two known functional domains: an N-terminal T hydroxylase (TH) homologous to the 5-methylcytosine hydroxylase domain in TET proteins and a J-DNA-binding domain (JDBD) that resides in the middle of JBP1. Here, we show that removing JDBD from JBP1 results in a soluble protein (Δ-JDBD) with the N- and C-terminal regions tightly associated together in a well-ordered structure. We found that this Δ-JDBD domain retains TH activity in vitro but displays a 15-fold lower apparent rate of hydroxylation compared with JBP1. Small-angle X-ray scattering (SAXS) experiments on JBP1 and JDBD in the presence or absence of J-DNA and on Δ-JDBD enabled us to generate low-resolution three-dimensional models. We conclude that Δ-JDBD, and not the N-terminal region of JBP1 alone, is a distinct folding unit. Our SAXS-based model supports the notion that binding of JDBD specifically to J-DNA can facilitate T hydroxylation 12-14 bp downstream on the complementary strand of the J-recognition site. We postulate that insertion of the JDBD module into the Δ-JDBD scaffold during evolution provided a mechanism that synergized J recognition and T hydroxylation, ensuring inheritance of base J in specific sequence patterns following DNA replication in kinetoplastid parasites. © 2019 Adamopoulos et al.

Citation

Athanassios Adamopoulos, Tatjana Heidebrecht, Jeroen Roosendaal, Wouter G Touw, Isabelle Q Phan, Jos Beijnen, Anastassis Perrakis. The domain architecture of the protozoan protein J-DNA-binding protein 1 suggests synergy between base J DNA binding and thymidine hydroxylase activity. The Journal of biological chemistry. 2019 Aug 23;294(34):12815-12825