Authors
Prem Shrestha, Sharon Lin, Ainsley Lederer, Nuwani Dharmapriya, Yasin Pourfarjam, Matthew Medearis, Angela Heintel, John A Tainer, Joel Moss, In-Kwon Kim
Published in
The Journal of biological chemistry. Pages 113695. Oct 26, 2026. Epub Oct 26, 2026.
Abstract
Serine MARylation (Ser-MARylation), catalyzed by poly(ADP-ribose) polymerase 1/2 (PARP1/2)-Histone PARylation factor 1 (HPF1) complex and reversed by ADP-ribosyl-acceptor hydrolase 3 (ARH3), is the primary form of DNA damage-induced ADP-ribosylation in human cells. Distinct from PARylation, Ser-MARylation represents a critical "second wave" of signaling following DNA damage. Dysregulation of Ser-MARylation is implicated in neurodegeneration and cancer, and ARH3 deficiency drives resistance to PARP inhibitor therapy. However, mechanistic characterization of Ser-MARylation reversal has been hampered by the lack of well-defined protein substrates and sensitive quantitative tools. Here, we report a quantitative assay to monitor ARH3-mediated reversal of protein-linked Ser-MARylation using Ser-MARylated HPF1 as a specific, defined substrate. Strikingly, ARH3 displays a greater than 200-fold kinetic preference for Ser-MARylated HPF1 over MARylated PARP1. By using a panel of ADP-ribose analogs, we identified ADP-HPD as an effective ARH3 inhibitor that mimics the oxocarbenium-like transition state of O-glycosidic bond hydrolysis. Together with structural and mutational analysis, these findings support a catalytic mechanism in which Asp77 activates a catalytic water for nucleophilic attack on the scissile serine-ADP-ribose bond. Collectively, this work establishes a mechanistic framework for Ser-MARylation reversal and lays the groundwork for rational ARH3 inhibitor development.
PMID:
42933574
Bibliographic data and abstract were imported from PubMed on 27 Oct 2026.
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