Synthesis of 5-fluoro-2'-deoxyuridine using recombinant Escherichia coli thymidine phosphorylase
2012
Beresnev, A.I. | Kucharskaya, T.A. | Kvach, S.V. | Zinchenko, A.I., National Academy of Sciences (Belarus). Institute of Microbiology
The present study was undertaken in order to develop practical syanthesis of 5-fluoro-2'-deoxyuridine using reaction of enzymatic transglycosylation. The first application of recombinant Escherichia coli thymidine phosphorylase to produce 5-fluoro-2'-deoxyuridine was demonstrated. In course of the study there was presented a scheme for synthesis of 5-fluoro-2'-deoxyuridine from 2'-deoxyuridine and 5-fluorouracil by recombinant TPase of E.coli. Also there were shown the effects of variation of fluorouracil and 2'-deoxyuridine concentrations on the substrate conversion with recombinant TPase. The molar ratio of substrates and concentration of phosphate ions in the reaction medium were shown to be significant factors affecting the desired product yield. Under optimal reaction conditions: 2 mM potassium phosphate buffer (pH 7.0); 10 mM 2'-deoxyuridine; 50 mM 5-fluorouracil; 1.5 units/ml of thymidine phosphorylase; incubation time 1r at temperature 40 deg C, the yield of 5-fluoro-2'-deoxyuridine reached more than 95% with respect to the initial 2'-deoxyuridine level. The obtained results open up possibilities for engineering of industrial technology for production of the title pharmaceutically valuable modified nucleeoside.
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