Selective probing of ADP-ribosylation reactions with oxidized 2'-deoxy-nicotinamide adenine dinucleotide.
A homogeneous preparation of an arginine-specific mono(ADP-ribosyl)transferase from turkey erythrocytes effectively utilized 2'-deoxy-NAD+ for the 2'-deoxy(ADP-ribose) modification of arginine methyl ester with an apparent Km of 27.2 microM and a Vmax of 36.4 mumol min-1 (mg of protein)-1. The adduct formed was also used as a substrate by an avian erythrocyte arginine(ADP-ribose)-specific hydrolase that generated free 2'-deoxy(ADP-ribose). In contrast, 2'-deoxy-NAD+ was not a substrate in the initiation or elongation reaction catalyzed by highly purified poly(ADP-ribose) polymerase from calf thymus. However, 2'-deoxy-NAD+ was a potent noncompetitive inhibitor of NAD+ in the elongation reaction catalyzed by the polymerase, with an apparent Ki of 32 microM. These results indicate that 2'-deoxy-NAD+ may be utilized to specifically identify protein acceptors for endogenous mono(ADP-ribosyl)transferases in complex biological systems that may contain a high activity of poly(ADP-ribose) polymerase, i.e., cell nuclei preparations.
D036002:ADP Ribose Transferases; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D004912:Erythrocytes / Q000201:enzymology; D007700:Kinetics; D009243:NAD / Q000031:analogs & derivatives* / Q000378:metabolism; D010084:Oxidation-Reduction; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism
D009243:NAD; D000246:Adenosine Diphosphate Ribose; C021004:2'-deoxynicotinamide adenine dinucleotide; D036002:ADP Ribose Transferases; D011065:Poly(ADP-ribose) Polymerases