Two-step efficient synthesis of 5-methyluridine via two thermostable nucleoside phosphorylase from Aeropyrum pernix

5-Methyluridine has been synthesized in high yield using guanosine and thymine as starting materials in the presence of highly thermostable recombinant purine nucleoside phosphorylase (PNP) and uridine phosphorylase (UP) obtained from hyperthermophilic aerobic crenarchaeon Aeropyrum pernix. Key reaction parameters such as pH, temperature, concentration of buffer and substrates were investigated. At the optimal conditions, 5-methyluridine was achieved in yield 85% with a guanosine conversion of 96% in 10ml scale. The process can be performed at high temperature, which will highly increase the solubility of substrates, therefore, this process is suitable for the industry application.