Amino acid specific ADP-ribosylation: substrate specificity of an ADP-ribosylarginine hydrolase from turkey erythrocytes.
An ADP-ribosylarginine hydrolase, which catalyzes the degradation of ADP-ribosyl[14C]arginine to ADP-ribose plus arginine, was separated by ion exchange, hydrophobic, and gel permation chromatography from NAD:arginine ADP-ribosyltransferases, which are responsible for the stereospecific formation of alpha-ADP-ribosylarginine. As determined by NMR, the specific substrate for the hydrolase was alpha-ADP-ribosylarginine, the product of the transferase reaction. The ADP-ribose moiety was critical for substrate recognition; (phosphoribosyl) [14C]arginine and ribosyl[14C]arginine were poor substrates and did not significantly inhibit ADP-ribosyl[14C]arginine degradation. In contrast, ADP-ribose was a potent inhibitor of the hydrolase and significantly more active than ADP greater than AMP greater than adenosine. In addition to ADP-ribosyl[14C]arginine, both ADP-ribosyl[14C]guanidine and (2'-phospho-ADP-ribosyl)[14C]arginine were also substrates; at pH greater than 7, ADP-ribosyl[14C]guanidine was degraded more readily than the [14C]arginine derivative. Neither arginine, guanidine, nor agmatine, an arginine analogue, was an effective hydrolase inhibitor. Thus, it appears that the ADP-ribosyl moiety but not the arginine group is critical for substrate recognition. Although the hydrolase requires thiol for activity, dithiothreitol accelerated loss of activity during incubation at 37 degrees C. Stability was enhanced by Mg2+, which is also necessary for optimal enzymatic activity. The findings in this paper are consistent with the conclusion that different enzymes catalyze ADP-ribosylarginine synthesis and degradation. Furthermore, since the hydrolase and transferases possess a compatible stereospecificity and substrate specificity, it would appear that the two enzymatic activities may serve as opposing arms in an ADP-ribosylation cycle.
D000227:Adenine Nucleotides / Q000494:pharmacology; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000818:Animals; D002250:Carbon Radioisotopes; D004912:Erythrocytes / Q000201:enzymology*; D006026:Glycoside Hydrolases; D006863:Hydrogen-Ion Concentration; D008190:Lyases / Q000097:blood*; D009699:N-Glycosyl Hydrolases; D013379:Substrate Specificity; D014422:Turkeys
D000227:Adenine Nucleotides; D002250:Carbon Radioisotopes; D000246:Adenosine Diphosphate Ribose; D006026:Glycoside Hydrolases; D009699:N-Glycosyl Hydrolases; C046422:ADP-ribosylarginine hydrolase; D008190:Lyases