CD38-mediated ribosylation of proteins.
The lymphocyte cell-surface Ag CD38 catabolizes NAD to adenosine 5' diphosphoribose (ADPR) and cyclic ADPR (cADPR). We show here that the soluble extracellular domain of CD38 (sCD38) mediates ADP ribosylation of several proteins. This was demonstrated by mass spectrometric analyses which revealed the addition of mass in units of 541.1 Da to these proteins, presumably corresponding to the covalent attachment of one or more ADPR moieties. Separate experiments showed that the same proteins became specifically radiolabeled following incubation with [32P]NAD plus sCD38. Additionally, it is shown that sCD38 can autoribosylate. Moreover, sCD38-mediated protein ribosylation was found to occur specifically at cysteine residues, since it was effectively blocked by addition of L-cysteine but not by other amino acids, and CD38-mediated protein ribosylation could be reversed by the addition of HgCl2, which specifically cleaves thiol-glycosidic bonds. ADPR purified from the reaction of sCD38 with NAD could itself be covalently transferred to target proteins at rates similar to the sCD38-mediated reaction, indicating that the ribosylation proceeds via the generation of this reactive intermediate. In vitro mutagenesis of a catalytic Glu residue that is conserved in numerous ADP-ribosyl transferases revealed that this amino acid is also important for catalysis in CD38. These data suggest that CD38 has the potential to cause ribosylation of experimental proteins, and raises the possibility that its specific ribosylation of a currently unidentified lymphocyte protein may contribute to its array of immunoregulatory activities.
Grimaldi
J C
JC
DNAX Research Institute of Molecular and Cellular Biology, Palo Alto, CA 94304, USA.
Balasubramanian
S
S
Kabra
N H
NH
Shanafelt
A
A
Bazan
J F
JF
Zurawski
G
G
Howard
M C
MC
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000246:Adenosine Diphosphate Ribose / Q000502:physiology; D000818:Animals; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D003545:Cysteine / Q000378:metabolism; D008562:Membrane Glycoproteins; D051379:Mice; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D011065:Poly(ADP-ribose) Polymerases / Q000502:physiology*; D011506:Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
D015703:Antigens, CD; D000943:Antigens, Differentiation; D008562:Membrane Glycoproteins; D011506:Proteins; D011994:Recombinant Proteins; D000246:Adenosine Diphosphate Ribose; D011065:Poly(ADP-ribose) Polymerases; D009699:N-Glycosyl Hydrolases; D036541:ADP-ribosyl Cyclase; C498155:Cd38 protein, mouse; D051997:ADP-ribosyl Cyclase 1; D003545:Cysteine