The role of uridine-cytidine kinase in the regulation of pyrimidine ribonucleotide synthesis in Tetrahymena pyriformis GL

Abstract

1. 1. Uridine—cytidine kinase (ATP:5′-uridine phosphotransferase, EC 2.7.1.48) has been purified 500-fold from Tetrahymena pyriformis GL, and some of its properties have been investigated. 2. 2. The molecular weight of the enzyme has been estimated as 195 000 by gel filtration. 3. 3. A vibalent cation is necessary for activity. MG 2+ is the most effective but may be partially replaced by Mn 2+ or Fe 2+. 4. 4. Uridine and cytidine are readily phosphorylated to UMP and CMP, respectively, however 2′-deoxyuridine, 2′-deoxycytidine, 5-methyluridine and 5-fluoro-2′-deoxyuridine do not serve as phosphate acceptors. 5. 5. The kinase utilizes ATP, dATP, GTP, dCTP and dUTP as phosphate donors at selected concentrations, while CTP, UTP and dGTP are not effective in this capacity. 6. 6. Substrate phosphorylation is inhibited by the addition of either UTP or CTP to the assay mixture. Inhibition by CTP is competitive with the phosphate donor and noncompetitive with the phosphate acceptor. 7. 7. It is suggested that UTP and CTP act as feedback inhibitors of uridine—cytidine kinase. Such control of pyrimidine ribonucleotide synthesis may be of importance in Tetrahymena, an organism which is unable to synthesize the pyrimidine ring by the de novo pathway.