Poly(ADP-ribosyl)ation which is mainly involved in DNA repair and replication is catalyzed mainly by poly(ADP-ribose) polymerase-1 (PARP-1) and poly(ADP-ribose) glycohydrolase (PARG) Although recombinant human PARP-1 (hPARP-1) is commercially available there are no reports on the preparation of recombinant human PARG (hPARG) Here we report the efficient expression and purification of a recombinant hPARG-catalytic domain (hPARG-CD) from Escherichia coil (E coil) hPARG-CD was expressed as a fusion protein with a glutathione S-transferase (GST) tag at the N-terminus and a hexahistidine (6His) tag at the C-terminus Both high cell density and low temperature culture conditions were important for the maximum production of soluble recombinant hPARG-CD After sequential affinity chromatography using immobilized metal affinity resin and glutathione-Sepharose (GSH-Sephasrose) more than 95% pure recombinant hPARG-CD was obtained with a yield of approximately 2 mg per 1 L of E colt culture medium The km and Vmax values of purified recombinant hPARG-CD were 9 0 mu M and 35 6 mu mol/min/mg protein respectively These kinetic values were similar to those of purified endogenous hPARG reported previously Furthermore the recombinant hPARG-CD was inhibited by known PARG inhibitors such as adenosine diphosphate (hydroxymethyl) pyrrolidinediol (ADP-HPD) eosin Y and phloxine B These results show that the recombinant hPARG-CD is useful to search for specific inhibitors and to elucidate the regulatory mechanisms of hPARG (C) 2010 Elsevier Inc All rights reserved