Hydrolysis of NADP+ by platelet CD38 in the absence of synthesis and degradation of cyclic ADP-ribose 2'-phosphate.
CD38 is a multifunctional cell surface ectoenzyme that catalyzes both the synthesis of cyclic ADP-ribose from NAD+ and its hydrolysis to ADP-ribose. In this work, we investigated the metabolism of NADP+ by CD38 expressed on human platelets. Incubation of either platelet membranes or intact cells with NADP+ resulted in the rapid and time-dependent accumulation of ADP-ribose 2'-phosphate that paralleled the consumption of the substrate. However, under the same conditions, synthesis of cyclic ADP-ribose 2'-phosphate was not observed. By immunoprecipitation experiments, we identified CD38 as the enzyme responsible for the observed NADP+ glycohydrolase activity. The lack of detection of cyclic ADP-ribose 2'-phosphate was not due to its rapid hydrolysis, since direct incubation of platelet membranes with cyclic ADP-ribose 2'-phosphate did not result in the formation of ADP-ribose 2'-phosphate. By contrast, the same membrane samples expressed a significant ability to hydrolyze cyclic ADP-ribose to ADP-ribose. The absence of cyclic ADP-ribose 2'-phosphate hydrolase activity was also confirmed using high concentrations of substrate and by analysing both intact Jurkat T-lymphocytes and immunoprecipitated CD38. These results indicate that CD38, which is a multifunctional enzyme towards NAD+, displays exclusively a NADP+ glycohydrolase activity and is unable to catalyze both the synthesis and the hydrolysis of cyclic ADP-ribose 2'-phosphate.
Torti
M
M
Department of Biochemistry, University of Pavia, Italy.
Bertoni
A
A
Canobbio
I
I
Sinigaglia
F
F
Balduini
C
C
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000378:metabolism*; D001792:Blood Platelets / Q000201:enzymology* / Q000276:immunology*; D036563:Cyclic ADP-Ribose / Q000031:analogs & derivatives*; D006801:Humans; D006868:Hydrolysis; D066298:In Vitro Techniques; D019169:Jurkat Cells; D007700:Kinetics; D008562:Membrane Glycoproteins; D009244:NAD+ Nucleosidase / Q000378:metabolism*; D009249:NADP / Q000378:metabolism*; D013379:Substrate Specificity; D013601:T-Lymphocytes / Q000378:metabolism
D015703:Antigens, CD; D000943:Antigens, Differentiation; D008562:Membrane Glycoproteins; C098663:cyclic ADP-ribose 2'-phosphate; D036563:Cyclic ADP-Ribose; C020857:phosphoadenosine diphosphoribose; D000246:Adenosine Diphosphate Ribose; D009249:NADP; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D009244:NAD+ Nucleosidase; D051997:ADP-ribosyl Cyclase 1