Enzymatic Synthesis Of Pyrimidine Nucleotides

Nucleotides and their derivatives have important application value in agriculture, medicine, food, genetic engineering and scientific research. A new synthesis method of 5’-UMP, 5’-CMP and 5’-UTP- was proposed in this study, which uridine-cytidine kinase(UCK), uridine monophosphate kinase(UMPK) and acetate kinase(ACK) were used. Gene uck, ack and pyrH were from E. coli, and gene uck1, uck2 and ack-BL were from Lactobacillus bulgaricus In the reaction of synthesis of 5’-UMP and 5’-CMP, when UCK from recombinant strain D28U and ACK from DA-L were used as biocatalyst, the yields of 5’-UMP and 5’-CMP were high. While uridine or cytidine was the substrate, the yields of 5’-UMP or 5’-CMP could reach 97%. The former reaction need 3 h to balance, and the later need 12 h or more. GTP was the optimal phosphate donor in the phosphorylation of uridine or cytidine. When GTP was 1/60 of the initial substrate concentration, it was enough to the phosphorylation reaction. In the synthesis reaction of 5’-UTP directly from uridine, uridine-cytidine kinase, acetate kinase and uridine monophosphate kinase added, GTP and ATP as the phosphate donors, the yield of 5’-UTP could attain 90% in 6 h.The synthesis method proposed in this study was not only simple and friendly environmental, but also low-cost, high conversion rates and no degradation of end products. These methods were very suitable for industrial production.