嘌呤核苷磷酸化酶
化学
核苷
酶
嘧啶
生物化学
嘌呤
作者
Immacolata Serra,Daniela Ubiali,Jure Piškur,Stig Christoffersen,Elizabeth Sandra Lewkowicz,Adolfo M. Iribarren,Alessandra M. Albertini,Marco Terreni
标识
DOI:10.1002/cplu.201200278
摘要
Abstract The use of nucleoside phosphorylases (NPs; EC 2.4.2.n) represents a convenient alternative to the chemical route for the synthesis of natural and modified nucleosides. We purified four recombinantly expressed nucleoside phosphorylases from the bacterial pathogens Citrobacter koseri , Clostridium perfringens , and Streptococcus pyogenes ( Ck PNPI, Ck PNPII, Cp UP, Sp UP) and their substrate specificity was investigated towards either natural pyrimidine or purine nucleosides and some analogues, namely, arabinosyladenine (araA) and 2′,3′‐dideoxyinosine (ddI). A 2–3 % activity towards these latter compounds (compared to the natural substrates) was observed. Enzyme activities were compared to the specificities obtained for the enzymes pyrimidine nucleoside phosphorylase from Bacillus subtilis ( Bs PyNP) and purine nucleoside phosphorylase from Aeromonas hydrophila ( Ah PNPII) previously reported by some of the authors. The enzymes displaying the suitable specificity for the synthesis of araA and ddI were immobilized on aldehyde–agarose. The immobilized preparations were highly stable at alkaline pH and in the presence of methanol or acetonitrile as cosolvent. They were used in the synthesis of araA and ddI by a one‐pot, bienzymatic transglycosylation achieving 74 and 44 % conversion, respectively.
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