Cloning and characterization of a novel membrane-associated lymphocyte NAD:arginine ADP-ribosyltransferase.
Mono-ADP-ribosylation is a post-translational modification of proteins in which the ADP-ribose moiety of NAD is transferred to proteins and is responsible for the toxicity of some bacterial toxins (e.g. cholera toxin and pertussis toxin). NAD:arginine ADP-ribosyltransferases cloned from human and rabbit skeletal muscle and from mouse lymphoma (Yac-1) cells are glycosylphosphatidylinositol-anchored and have similar enzymatic and physical properties; transferases cloned from chicken heterophils and red cells have signal peptides and may be secreted. We report here the cloning and characterization of an ADP-ribosyltransferase (Yac-2), also from Yac-1 lymphoma cells, that differs in properties from the previously identified eukaryotic transferases. The nucleotide and deduced amino acid sequences of the Yac-1 and Yac-2 transferases are 58 and 33% identical, respectively. The Yac-2 protein is membrane-bound but, unlike the Yac-1 enzyme, appears not to be glycosylphosphatidylinositol-anchored. The Yac-1 and Yac-2 enzymes, expressed as glutathione S-transferase fusion proteins in Escherichia coli, were used to compare their ADP-ribosyltransferase and NAD glycohydrolase activities. Using agmatine as the ADP-ribose acceptor, the Yac-1 enzyme was predominantly an ADP-ribosyltransferase, whereas the transferase and NAD glycohydrolase activities of the recombinant Yac-2 protein were equivalent. The deduced amino acid sequence of the Yac-2 transferase contained consensus regions common to several bacterial toxin and mammalian transferases and NAD glycohydrolases, consistent with the hypothesis that there is a common mechanism of NAD binding and catalysis among ADP-ribosyltransferases.
Okazaki
I J
IJ
Pulmonary-Critical Care Medicine Branch, NHLBI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Kim
H J
HJ
Moss
J
J
D036002:ADP Ribose Transferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002462:Cell Membrane / Q000201:enzymology; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry; D006801:Humans; D008214:Lymphocytes / Q000201:enzymology*; D051379:Mice; D008969:Molecular Sequence Data; D009244:NAD+ Nucleosidase / Q000378:metabolism; D011817:Rabbits; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D036002:ADP Ribose Transferases; D009244:NAD+ Nucleosidase