Molecular characterization of rat T lymphocyte alloantigen RT6.1 as an ADP-ribosyltransferase.
A family of glycosylphosphatidylinositol-linked ADP-ribosyltransferases, of which cDNAs were cloned from various mammalian cells, possess a common Glu-rich motif (EEEVLIP) near their carboxyl termini. Although the first Glu in the common motif is replaced by Gln (Q207EEVLIP) in rat T lymphocyte alloantigens RT6.1 and RT6.2, the two RT6s appear to have ADP-ribosyltransferase activity. To investigate the significance of the Glu-rich motif in the enzyme activity, we produced a mutant RT6.1, in which Gln207 was replaced by Glu (Q207E), together with wild-type RT6s, in Escherichia coli. The recombinant RT6.1 and RT6.2 displayed extremely low auto-ADP-ribosylation, though the latter modification was somewhat higher than the former one. In contrast, much higher the auto-modification was observed for Q207E mutant. Moreover, the mutant could effectively ADP-ribosylate agmatine as a substrate. Thus, the single amino acid mutation of RT6.1 caused remarkable increase in its ADP-ribosyltransferase activity, indicating that the Glu-rich motif near the carboxy terminus plays an important role in the enzyme activity.
Maehama
T
T
Department of Physiological Chemistry, Faculty of Pharmaceutical Sciences, University of Tokyo, Japan.
Katada
T
T
D036002:ADP Ribose Transferases; D000818:Animals; D000945:Antigens, Differentiation, T-Lymphocyte; D058851:GPI-Linked Proteins; D018698:Glutamic Acid; D006649:Histocompatibility Antigens / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008562:Membrane Glycoproteins; D016297:Mutagenesis, Site-Directed; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics; D013601:T-Lymphocytes / Q000201:enzymology*
D000945:Antigens, Differentiation, T-Lymphocyte; D058851:GPI-Linked Proteins; D006649:Histocompatibility Antigens; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D018698:Glutamic Acid; D036002:ADP Ribose Transferases; D011065:Poly(ADP-ribose) Polymerases; C449301:ART1 protein, human; C090163:Art2b protein, rat