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A comparison between immobilized pyrimidine nucleoside phosphorylase from Bacillus subtilis and thymidine phosphorylase from Escherichia coli in the synthesis of 5-substituted pyrimidine 2′-deoxyribonucleosides

Serra, Immacolata, Bavaro, Teodora, Cecchini, Davide A., Daly, Simona, Albertini, Alessandra M., Terreni, Marco, Ubiali, Daniela
Journal of Molecular Catalysis. B, Enzymatic 2013 v.95 pp. 16-22
Bacillus subtilis, Escherichia coli, biocatalysts, biotransformation, catalytic activity, immobilized enzymes, pH, phosphates, phosphorylase, substrate specificity, thymidine
Pyrimidine nucleoside phosphorylase from Bacillus subtilis (BsPyNP, E.C. and thymidine phosphorylase from Escherichia coli (EcTP, E.C. were used, as immobilized enzymes, in the synthesis of 5-halogenated pyrimidine 2′-deoxyribonucleosides (14–18) by transglycosylation in fully aqueous medium. From the comparative study of the two biocatalysts, no remarkable differences emerged about their substrate specificity, bioconversion yield, stability in organic cosolvents (DMF and MeCN). Moreover, both biocatalysts could be recycled for at least 5 times with no loss of the productivity.Both enzymes do not accept arabinonucleosides and 2′,3′-dideoxynucleosides as substrates, whereas they catalyze bioconversions involving 5′-deoxyribonucleosides and 5-halogenated uracils. The synthesis of compounds 14–18 proceeded at a similar conversion (33–68% for BsPyNP and 25–62% for EcTP, respectively). Immobilization was found to exert, for both the biocatalysts, a dramatic enhancement of stability upon incubation in MeCN.Optimization of 5-fluoro-2′-deoxyuridine (14) synthesis (pH 7.5, 10mM phosphate buffer, nucleoside/nucleobase 3:1 molar ratio) and subsequent scale-up afforded the target compound in 73% (EcTP) or 76% (BsPyNP) conversion (about 9g/L).