9tru | pdb_00009tru
Zebrafish dUTPase in complex with staphylococcal Stl repressor
Structural highlights
FunctionQ5XJ23_DANRE Catalyzes the cleavage of 2'-deoxyuridine 5'-triphosphate (dUTP) into 2'-deoxyuridine 5'-monophosphate (dUMP) and inorganic pyrophosphate and through its action efficiently prevents uracil misincorporation into DNA and at the same time provides dUMP, the substrate for de novo thymidylate biosynthesis. Inhibits peroxisome proliferator-activated receptor (PPAR) activity by binding of its N-terminal to PPAR, preventing the latter's dimerization with retinoid X receptor. Essential for embryonic development.[ARBA:ARBA00057946] Involved in nucleotide metabolism via production of dUMP, the immediate precursor of thymidine nucleotides, and decreases the intracellular concentration of dUTP so that uracil cannot be incorporated into DNA.[RuleBase:RU367024] Publication Abstract from PubMedDeoxyuridine triphosphatase (dUTPase) is essential for DNA replication fidelity and embryonic development in metazoa, yet whether a protein inhibitor can acutely disrupt dUTPase function in a living vertebrate has not been tested. The Staphylococcus aureus Stl protein inhibits diverse dUTPases in vitro, but its efficacy in a eukaryotic organism was unknown. We determined the crystal structure of zebrafish dUTPase in complex with the N-terminal Stl fragment to 2.26 A resolution, revealing conserved active-site engagement and displacement of the C-terminal arm. Biolayer interferometry confirmed low-nanomolar binding affinity with K(D) = 0.7 nM, and enzyme assays demonstrated ~50% inhibition of zebrafish dUTPase activity at saturating Stl concentrations. These structural and biophysical data establish that Stl forms a tight inhibitory complex with zebrafish dUTPase, prompting us to test whether this interaction can perturb enzyme function in vivo. Microinjection of Stl or its point mutant, retaining dUTPase-inhibitory activity but unable to bind DNA, into fertilized zebrafish oocytes reduces embryo survival to ~60% within 48 h compared to ~90% in controls, demonstrating that lethality depends on dUTPase targeting rather than DNA binding. Accelerated mortality relative to genetic dUTPase knockout indicates that maternally deposited dUTPase supports viability during early cleavage. Stl thus acts as an immediate, proteinaceous dUTPase inhibitor in a living vertebrate, providing a tool to dissect dUTPase function and regulation during embryonic development. A bacterial protein inhibitor of dUTPase disrupts zebrafish embryogenesis through a conserved active-site mechanism.,Perey-Simon V, Toth ZS, Dombovari D, Kazzazy L, Varga M, Vertessy BG, Nagy GN, Nyiri K Protein Sci. 2026 Oct;35(10):e70767. doi: 10.1002/pro.70767. PMID:42747148[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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