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Molecular basis for DarT ADP-ribosylation of a DNA base

DOI: 10.1038/s41586-021-03825-4 DOI Help

Authors: Marion Schuller (University of Oxford) , Rachel E. Butler (University of Surrey) , Antonio Ariza (University of Oxford) , Callum Tromans-Coia (University of Oxford) , Gytis Jankevicius (University of Oxford; University of Basel) , Tim D. W. Claridge (University of Oxford) , Sharon L. Kendall (The Royal Veterinary College) , Shan Goh (The Royal Veterinary College) , Graham R. Stewart (University of Surrey) , Ivan Ahel (University of Oxford)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Nature , VOL 58

State: Published (Approved)
Published: August 2021
Diamond Proposal Number(s): 18069

Open Access Open Access

Abstract: ADP-ribosyltransferases use NAD+ to catalyse substrate ADP-ribosylation1, and thereby regulate cellular pathways or contribute to toxin-mediated pathogenicity of bacteria2,3,4. Reversible ADP-ribosylation has traditionally been considered a protein-specific modification5, but recent in vitro studies have suggested nucleic acids as targets6,7,8,9. Here we present evidence that specific, reversible ADP-ribosylation of DNA on thymidine bases occurs in cellulo through the DarT–DarG toxin–antitoxin system, which is found in a variety of bacteria (including global pathogens such as Mycobacterium tuberculosis, enteropathogenic Escherichia coli and Pseudomonas aeruginosa)10. We report the structure of DarT, which identifies this protein as a diverged member of the PARP family. We provide a set of high-resolution structures of this enzyme in ligand-free and pre- and post-reaction states, which reveals a specialized mechanism of catalysis that includes a key active-site arginine that extends the canonical ADP-ribosyltransferase toolkit. Comparison with PARP–HPF1, a well-established DNA repair protein ADP-ribosylation complex, offers insights into how the DarT class of ADP-ribosyltransferases evolved into specific DNA-modifying enzymes. Together, our structural and mechanistic data provide details of this PARP family member and contribute to a fundamental understanding of the ADP-ribosylation of nucleic acids. We also show that thymine-linked ADP-ribose DNA adducts reversed by DarG antitoxin (functioning as a noncanonical DNA repair factor) are used not only for targeted DNA damage to induce toxicity, but also as a signalling strategy for cellular processes. Using M. tuberculosis as an exemplar, we show that DarT–DarG regulates growth by ADP-ribosylation of DNA at the origin of chromosome replication.

Journal Keywords: DNA damage and repair; DNA repair enzymes; Pathogens

Diamond Keywords: Bacteria; Tuberculosis (TB)

Subject Areas: Biology and Bio-materials, Chemistry

Instruments: I03-Macromolecular Crystallography , I04-1-Macromolecular Crystallography (fixed wavelength) , I04-Macromolecular Crystallography , I24-Microfocus Macromolecular Crystallography

Added On: 24/08/2021 10:10

Discipline Tags:

Pathogens Infectious Diseases Health & Wellbeing Biochemistry Genetics Chemistry Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)