Glutamic Acid 207 in Rodent T-cell RT6 Antigens Is Essential for Arginine-specific ADP-ribosylation

• Published in: The Journal of biological chemistry 1996-11, Vol.271 (47), p.29552-29555
• Main Authors: Hara, Nobumasa, Tsuchiya, Mikako, Shimoyama, Makoto
• Format: Article
• Language: English
• Subjects: Adenosine Diphosphate Ribose - metabolism; ADP Ribose Transferases; Amino Acid Sequence; Animals; Antigens, Differentiation, T-Lymphocyte; Arginine - metabolism; Glutamic Acid - metabolism; Histocompatibility Antigens - metabolism; Membrane Glycoproteins; Mice; Molecular Sequence Data; Mutagenesis; Rats; Sequence Homology, Amino Acid; T-Lymphocytes - immunology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.271.47.29552

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 Gln at position 207 in RT6.1, we replaced the Gln with Glu, Asp, or Ala, 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 kcat of the Glu-207 mutant increased compared to findings with the Asp-207 mutant. Moreover, the mouse homologue of rat RT6 lost arginine-specific ADP-ribosyltransferase activity when Glu-207 was replaced with Gln. Thus, Glu-207 in rodent T-cell RT6 antigens is essential for transfer reaction of ADP-ribose to arginine.