Phosphorylation of Nucleotides with Inorganic Cyclo-Triphosphate

Article abstract of DOI:10.1246/bcsj.64.490

Phosphorylation of nucleotides (nucleoside 3'- and 5'-monophosphates, and 2'-deoxynucleoside 5'-monophosphates) with inorganic sodium cyclotriphosphate (P_3m) was studied in aqueous solutions under various conditions (mixing ratio of P_3m to nucleotides, pH, reaction temperature, and time). (1) Unprotected nucleoside 5'-monophosphates (5'-NMP's) were easily phosphorylated at the cis-2',3'-diol by P_3m to form selectively nucleoside 2',5'-bis(monophosphate) (2',5'-NDP's), nucleoside 3',5'-bis(monophosphate) (3',5'-NDP's), and nucleoside 2',3'-cyclic 5'-bis(monophosphate) (cNDP's). (2) The phosphorylation of 5'-NMP's was strongly dependent on mixing ratio, pH, reaction temperature, and time.Under conditions of high mixing ratios of P_3m to 5'-NMP's (5:1 - 10:1), high pH (12), and room temperature, 92 -98percent of 5'-NMp's was converted into 3',5'-NDP's and 2',5'-NDP's in roughly equimolar quantities. (3) Small quantities (5 -8percent) of cNdP's were formed at the initial stage of reaction of 5'-NMP's with P_3m but in the course of the reaction for a long period, cNDP's were hydrolyzed to 2',5'-NDP's and 3',5'-NDP's. (4) Nucleoside 3'-monophosphates (3'-NMP's) and 2'-deoxynucleoside 5'-monophosphates (dNMP's) could not be phosphorylated by P_3m, which indicates that the presence of hydroxyl groups at both 2'- and 3'-positions on nucleotides is indispensable for the phosphorylation of nucleotides with P_3m. (5) The mechanism of the formation of 2',5'-NDP's, 3',5'-NDP's, and cNDP's in the phosphorylation of 5'-NMP's with P_3m is discussed.