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

Abstract

Proteins can be reversibly modified through the addition of repeating, polymerized ADP-ribose (PAR) subunits catalysed by poly(ADP-ribose) polymerase (PARP). Removal of PAR requires a glycohydrolase (PARG), which cleaves the ribose–ribose bond between subunits. Ivan Ahel and colleagues report that bacteria and fungi have a divergent PARG, which is unrelated to other enzymes that cleave PAR. Its structure, in complex with ADP-ribose and with a PARG inhibitor, and the results of mutational analysis suggest that the mechanism used in mammals and bacteria may be conserved. PARP inhibitors are being developed as pharmaceuticals for diseases including cancer, and this work suggests that small, cell-permeable PARG inhibitors might also be possible drug candidates. 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.