Glutamic acid 207 in rodent T-cell RT6 antigens is essential for arginine-specific ADP-ribosylation

A rat T-cell antigen RT6.1 catalyzes NAD glycohydrolysis but not ADP-ribose transfer, even though the antigen has significant amino acid identity with eucaryotic arginine-specific ADP-ribosyltransferases. Since a highly conserved Glu in the catalytic region of these transferases is substituted with Gin at position 207 in RT6.1, we replaced the Gin with Glu, Asp, or Ale, by site-directed mutagenesis. The Glu-207 mutant produced ADP-ribosylarginine during incubation with NAD and L-arginine, The Asp-207 mutant but not the Ala 207 mutant produced ADP-ribosylarginine, but at a lower rate. In contrast, these mutations affected NAD glycohydrolase activity of RT6.1 to a much lesser extent. Kinetic studies of transferase reaction revealed that k(cat) of the Glu-207 mutant increased compared to findings with the Asp-207 mutant. Moreover, the mouse homologue of rat RT6 last arginine-specific ADP ribosyltransferase activity when Glu-207 was replaced with Grin. Thus, Glu-207 in rodent T cell RT6 antigens is essential for transfer reaction of ADP-ribose to arginine.