CHARACTERIZATION OF MOUSE RT6.1 NAD-ARGININE ADP-RIBOSYLTRANSFERASE

Rat RT6 proteins, and perhaps mouse Rt6, identify a set of immunoregul atory T lymphocytes. Rat RT6.1 (RT6.1) and rat RT6.2 (RT6.2) are NAD g lycohydrolases, which catalyze auto-ADP-ribosylation, but not ADP-ribo sylation of exogenous proteins. Mouse Rt6.1 (mRt6.1) also catalyzes au to-ADP-ribosylation. The activity of mouse cytotoxic T lymphocytes is reportedly inhibited by ADP-ribosylation of surface proteins, raising the possibility that mRt6 may participate in this process. The reactio ns catalyzed by mRt6, would, however, need to be more diverse than tho se of the rat homologues and include the ADP-ribosylation of accepters other than itself. To test this hypothesis, mRt6.1 and rat RT6.2 were synthesized in Sf9 insect cells and rat mammary adenocarcinoma (NMU) cells. mRt6.1, but not rat RT6.2, catalyzed the ADP-ribosylation of gu anidino-containing compounds (e.g. agmatine). Unlike RT6.2, mRt6.1 was a weak NAD glycohydrolase. In the presence of agmatine, however, the ratio of [adenine-C-14]ADP-ribosylagmatine formation from [adenine-C-1 4]NAD to [carbonyl-C-14]nicotinamide formation from [carbonyl-C-14]NAD was similar to 1.0, demonstrating that mRtG.1 is primarily a transfer ase. ADP-ribosylarginine hydrolase, which preferentially hydrolyzes th e alpha-anomer of ADP-ribosylarginine, released [U-C-14]arginine from ADP-ribosyl[U-C-14] arginine synthesized by mRT6.1, consistent with th e conclusion that mRt6.1 catalyzes a stereospecific S(n)2-like reactio n. Thus, mRt6.1 is an NAD:arginine ADP-ribosyltransferase capable of c atalyzing a multiple turnover, stereospecific S(n)2-like reaction.