Nucleoside 5'-diphosphates as effectors of mammalian ribonucleotide reductase

It was found that nucleoside 5'-diphosphates could serve as effectors of ribonucleotide reductase. ADP was an activator of CDP reduction; ADP reduction was activated by dGDP; GDP reduction was activated by dTDP. Conversely, dADP inhibited the reduction of CDP, UDP, GDP, and ADP; dGDP inhibited UDP and GDP reductions; and dTDP inhibited UDP reduction. The inhibition of UDP reduction by dADP, dTDP, and dGDP was at least equal to that observed for dATP, dTTP, and dGTP, respectively. In these experiments with the nucleoside diphosphates as effectors, high-pressure liquid chromatography analysis of the reaction mixtures showed that no nucleoside 5'-triphosphates were found during the reaction period which could account for the effects seen with the nucleoside diphosphates as effectors. Further experiments were carried out in which adenyl-5'-yl imidodiphosphate was used as the positive effector of CDP and UDP reductions in place of ATP. Under these conditions, CDP and UDP reductions were inhibited by dADP, dTDP, and dGDP to the same extent observed in the presence of ATP. ADP served not only as a substrate for ribonucleotide reductase but also as an activator of CDP and UDP reductions. The direct products (dNDPs) also served as positive and negative effectors. Dixon plots indicated that the dNDPs were acting as noncompetitive inhibitors with respect to the substrate. ADP increased the sedimentation velocity of the ribonucleotide reductase in a manner similar to ATP. These data are consistent with the allosteric effects seen with the nucleoside 5'-triphosphates. Additionally, from the thorough study of the role of effectors on UDP reduction, it is clear that UDP reduction was most sensitive to the negative effectors dATP, dADP, dTTP, dTDP, dGTP, and dGDP.