Function of NAD glycohydrolase in ADP-ribose uptake from NAD by human erythrocytes.
The function of the ectoenzyme NAD glycohydrolase (NADase) in ADP-ribose uptake from extracellular NAD was studied in human erythrocytes that express relatively high NADase activity (adult erythrocytes) and erythrocytes expressing very low activity (newborn erythrocytes). The rates of ADP-ribose uptake from NAD in human erythrocytes were correlated with their NADase activities. In contrast, there was no significant difference in the rates of ADP-ribose uptake among these cells when incubated with ADP-ribose. These results indicate that ecto-NADase may have a role as supplier of ADP-ribose for its uptake into the cells and that the cleavage of NAD by NADase is necessary for the ADP-ribose uptake by human erythrocytes. From ADP-ribose uptake studies at 37 degrees C a Km of 0.7 +/- 0.05 microM and a Vmax of 2.04 +/- 0.1 pmol/min per microliter cell water was found for the uptake of [3H]ADP-ribose. The thiol-reactive reagents p-chloromercuribenzene sulfonic acid and N-ethylmaleimide inhibited the uptake ADP-ribose with IC50 values of 50 +/- 4 and 750 +/- 25 mM, respectively. Since efflux of [3H]ADP-ribose was negligible, the ADP-ribose transport system appears to be unidirectional. The unidirectionality was supported by the evidence that transported ADP-ribose was rapidly degraded to AMP which is impermeable to the membrane.
Kim
U H
UH
Department of Biochemistry, Chonbuk National University Medical School, Chonju, South Korea.
Han
M K
MK
Park
B H
BH
Kim
H R
HR
An
N H
NH
D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives / Q000097:blood* / Q000494:pharmacology; D000328:Adult; D000375:Aging / Q000097:blood; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D004912:Erythrocytes / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007231:Infant, Newborn; D009243:NAD / Q000097:blood*; D009244:NAD+ Nucleosidase / Q000037:antagonists & inhibitors / Q000097:blood*
D009243:NAD; D000246:Adenosine Diphosphate Ribose; D009244:NAD+ Nucleosidase