Effect of Overexpression of Purine Nucleoside Phosphorylase on Ribavirin Production from Inosine Producing Strain

Purine nucleoside phosphorylase(PNP)genes deo D(PNP702)and pup G(PNP816)were amplified from Bacillus subtilis 168,genome through polymerase chain reaction. Inosine producing strain B.,subtilis Q60,was used as the starting strain. Four recombined strains were constructed by overexpression of intracellular and extracellular secretion of PNP. By adding 1,H-1,2,4-triazole-3-carboxamide(TCA)into the culture medium,the recombined strains could be used for ribavirin synthesis. The results showed that the production of ribavirin through intracellular overexpression of PNP702,and PNP816 of the recombined strains was(8.33±0.57) g/L and(11.00±0.38) g/L,and that from the extracellular secretion overexpression of PNP702 and PNP816 of the recombined strains was(9.06±0.17) g/L and(12.67±0.60) g/L. The catalytic efficiency of PNP816 was higher than that of PNP702,and extracellular secretion was better than intracellular overexpression. The titer of ribavirin of Q60/p BSApup G,in which PNP816 was overexpressed extracellularly,was up to(12.67±0.60) g/L,resulting in a 5.3 folds enhancement compared with the control strain.