Identification and characterization of a mammalian 39-kDa poly(ADP-ribose) glycohydrolase.
ADP-ribosylation is a post-translational modification resulting from transfer of the ADP-ribose moiety of NAD to protein. Mammalian cells contain mono-ADP-ribosyltransferases that catalyze the formation of ADP-ribose-(arginine) protein, which can be cleaved by a 39-kDa ADP-ribose-(arginine) protein hydrolase (ARH1), resulting in release of free ADP-ribose and regeneration of unmodified protein. Enzymes involved in poly(ADP-ribosylation) participate in several critical physiological processes, including DNA repair, cellular differentiation, and carcinogenesis. Multiple poly(ADP-ribose) polymerases have been identified in the human genome, but there is only one known poly(ADP-ribose) glycohydrolase (PARG), a 111-kDa protein that degrades the (ADP-ribose) polymer to ADP-ribose. We report here the identification of an ARH1-like protein, termed poly(ADP-ribose) hydrolase or ARH3, which exhibited PARG activity, generating ADP-ribose from poly-(ADP-ribose), but did not hydrolyze ADP-ribose-arginine, -cysteine, -diphthamide, or -asparagine bonds. The 39-kDa ARH3 shares amino acid sequence identity with both ARH1 and the catalytic domain of PARG. ARH3 activity, like that of ARH1, was enhanced by Mg(2+). Critical vicinal acidic amino acids in ARH3, identified by mutagenesis (Asp(77) and Asp(78)), are located in a region similar to that required for activity in ARH1 but different from the location of the critical vicinal glutamates in the PARG catalytic site. All findings are consistent with the conclusion that ARH3 has PARG activity but is structurally unrelated to PARG.
Oka
Shunya
S
Pulmonary-Critical Care Medicine Branch, NHLBI, National Institutes of Health, Bethesda, MD 20892-1590, USA.
Kato
Jiro
J
Moss
Joel
J
D000595:Amino Acid Sequence; D000818:Animals; D001345:Autoradiography; D015153:Blotting, Western; D020134:Catalytic Domain; D002417:Cattle; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D017931:DNA Primers / Q000737:chemistry; D004229:Dithiothreitol / Q000494:pharmacology; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D006868:Hydrolysis; D008274:Magnesium / Q000737:chemistry; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D009699:N-Glycosyl Hydrolases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D009693:Nucleic Acid Hybridization; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D012266:Ribose / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013950:Thymus Gland / Q000378:metabolism; D013997:Time Factors
D017931:DNA Primers; D012333:RNA, Messenger; D011994:Recombinant Proteins; D012266:Ribose; D011065:Poly(ADP-ribose) Polymerases; D006026:Glycoside Hydrolases; C508145:ADPRS protein, human; C022407:poly ADP-ribose glycohydrolase; D009699:N-Glycosyl Hydrolases; C000599892:ARH1 protein, mouse; D008274:Magnesium; D004229:Dithiothreitol