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The structure and catalytic mechanism of a poly(ADP-ribose) glycohydrolase

Nature volume 477, pages 616–620 (2011)Cite this article

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Abstract

Post-translational modification of proteins by poly(ADP-ribosyl)ation regulates many cellular pathways that are critical for genome stability, including DNA repair, chromatin structure, mitosis and apoptosis1. Poly(ADP-ribose) (PAR) is composed of repeating ADP-ribose units linked via a unique glycosidic ribose–ribose bond, and is synthesized from NAD by PAR polymerases1,2. PAR glycohydrolase (PARG) is the only protein capable of specific hydrolysis of the ribose–ribose bonds present in PAR chains; its deficiency leads to cell death3,4. Here we show that filamentous fungi and a number of bacteria possess a divergent form of PARG that has all the main characteristics of the human PARG enzyme. We present the first PARG crystal structure (derived from the bacterium Thermomonospora curvata), which reveals that the PARG catalytic domain is a distant member of the ubiquitous ADP-ribose-binding macrodomain family5,6. High-resolution structures of T. curvata PARG in complexes with ADP-ribose and the PARG inhibitor ADP-HPD, complemented by biochemical studies, allow us to propose a model for PAR binding and catalysis by PARG. The insights into the PARG structure and catalytic mechanism should greatly improve our understanding of how PARG activity controls reversible protein poly(ADP-ribosyl)ation and potentially of how the defects in this regulation are linked to human disease.

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Figure 1: Phylogeny and functional relationship between DUF2263 and canonical-type PARGs.
Figure 2: PAR hydrolytic activities of divergent and canonical PARGs.
Figure 3: T. curvata PARG crystal structure in complex with ADP-ribose.
Figure 4: Structural basis of PAR glycohydrolysis.

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Protein Data Bank

Data deposits

Atomic coordinates and structure factors have been deposited with the Protein Data Bank under accession codes 3SIG, 3SIH, 3SII and 3SIJ.

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Acknowledgements

We thank G. Clark for genomic DNA from E. dispar, G. Smith for the PARP inhibitor, M. Rossi for purified proteins and R. Thorough for editing English. We thank D. Ahel, A. Jordan, D. Ogilvie, S. Terzic, D. Neuhaus and S. Eustermann for helpful discussions. We are grateful to B. Lüscher for the gift of the PARP10 expression plasmid, and K. Labib and the members of his laboratory for advice with yeast work. This work was funded by Cancer Research UK. D.S. holds an AXA Research Fund post-doctoral fellowship. D.L. is a Royal Society University Research Fellow. Access to Diamond beamlines is gratefully acknowledged.

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Author notes

  1. Dea Slade and Mark S. Dunstan: These authors contributed equally to this work.

Authors and Affiliations

  1. Cancer Research UK, Paterson Institute for Cancer Research, University of Manchester, Wilmslow Road, Manchester M20 4BX, UK

    Dea Slade, Eva Barkauskaite, Ria Weston & Ivan Ahel

  2. Université de Paris-Descartes, Faculté de Médecine, INSERM U1001, 156 rue de Vaugirard, 75015 Paris, France

    Dea Slade

  3. Manchester Interdisciplinary Biocentre, Princess Street 131, Manchester M1 7DN, UK

    Mark S. Dunstan, Neil Dixon & David Leys

  4. ICOA – UMR CNRS 6005 Université d’Orléans, Rue de Chartres, F-45067 Orléans, France

    Pierre Lafite

  5. Rudjer Boskovic Institute, Bijenicka 54, HR-10000 Zagreb, Croatia

    Marijan Ahel

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Contributions

D.S. performed biochemical and in vivo experiments, prepared proteins for crystallization, analysed data and wrote the manuscript. M.S.D. performed structural/biophysical studies and analysed data. E.B. performed biochemical and in vivo experiments; R.W. performed supporting studies. M.A. and N.D. performed LC/MS analyses; P.L. performed molecular modelling studies. I.A. and D.L. wrote the manuscript, designed experiments and analysed data.

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Correspondence to Ivan Ahel.

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Slade, D., Dunstan, M., Barkauskaite, E. et al. The structure and catalytic mechanism of a poly(ADP-ribose) glycohydrolase. Nature 477, 616–620 (2011). https://doi.org/10.1038/nature10404

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