| Abstract: | Cladribine (2-chloro-2'-deoxyadenosine) was synthesized using intact cells of the recombinant Escherichia coli strain producing Geobacillus stearothermophilus B-2194 thermostable purine-nucleoside phosphorylase II (EC 2.4.2.1). Use of the cells containing this thermostable enzyme allowed the process to be conducted at a temperature of 70 degrees C, which provided the maximal concentrations of sparingly soluble substrates. The best results were obtained with 2-chloroadenine as a modified base. The highest yield of the target 2-chloro-2'-deoxyadenosine (up to 95% in the case of deoxyguanosine) was reached when using 2'-deoxypurines as donors of deoxyribose. Use of thymidine for these purposes required its considerable molar excess over 2-chloroadenine (up to 6 : 1), which is connected with a nonoptimal amount of endogenous thymidine phosphorylase, necessary for synthesis of deoxyribose-1-phosphate, in the transglycosylation reaction. |
