Enzymatic synthesis of nucleosides by nucleoside phosphorylase co-expressed in Escherichia coli.
Nucleoside phosphorylase is an important enzyme involved in the biosynthesis of nucleosides. In this study, purine nucleoside phosphorylase and pyrimidine nucleoside phosphorylase were co-expressed in Escherichia coli and the intact cells were used as a catalyst for the biosynthesis of nucleosides. For protein induction, lactose was used in place of isopropyl β-D-1-thiogalactopyranoside (IPTG). When the concentration of lactose was above 0.5 mmol/L, the ability to induce protein expression was similar to that of IPTG. We determined that the reaction conditions of four bacterial strains co-expressing these genes (TUD, TAD, DUD, and DAD) were similar for the biosyntheses of 2,6-diaminopurine nucleoside and 2,6-diaminopurine deoxynucleoside. When the substrate concentration was 30 mmol/L and 0.5% of the recombinant bacterial cell volume was used as the catalyst (pH 7.5), a greater than 90% conversion yield was reached after a 2-h incubation at 50 °C. In addition, several other nucleosides and nucleoside derivatives were efficiently synthesized using bacterial strains co-expressing these recombinant enzymes.
State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, China. bioxianleid@163.com
Qing-bao
QB
Ding
Ling
L
Ou
Dong-zhi
DZ
Wei
Xiao-kun
XK
Wei
Yan-mei
YM
Xu
Chun-yan
CY
Zhang
D004926:Escherichia coli / Q000502:physiology*; D009705:Nucleosides / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D010430:Pentosyltransferases / Q000502:physiology*; D015202:Protein Engineering / Q000379:methods*
D009705:Nucleosides; D010430:Pentosyltransferases; C067995:nucleoside phosphorylase