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KMID : 1094720150200010037
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Biotechnology and Bioprocess Engineering
2015 Volume.20 No. 1 p.37 ~ p.43
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One-pot process of 2′-deoxyguanylic acid catalyzed by a multi-enzyme system
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Li Yanyu
Ding Qingbao
Ou Ling
Qian Yahui
Zhang Jiao
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Abstract
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2′-Deoxyguanylic acid (deoxyguanosine-5′-monophosphate, dGMP) is a substance required by living cells that is used extensively in reagents, fine chemicals and other industrial fields. Traditionally, dGMP is separated from DNA degradation products, which is low-yielding and time-consuming. Herein, we investigated a novel, one-pot multi-enzymatic cascade reaction to produce dGMP. This reaction involved purine nucleoside phosphorylase (PNPase) and acetate kinase (ACKase) from Escherichia coli, N-deoxyribosytransferase II (NDT-II) from Lactobacillus delbrueckii and deoxyguanosine kinase (dGKase) from Bacillus subtilis. During the reaction, the initial guanosine substrate was cleaved into guanine and ribose-1-phosphate by PNPase. Then, deoxyguanosine (dGR) was subsequently produced from a reaction between guanine and thymidine catalysed by NDT-II. Finally, the intermediate dGR was phosphorylated to dGMP by dGKase and a cytidine triphosphate (CTP) regeneration system that utilised acetyl phosphate via ACKase. A very small amount of CTP was added because CTP regeneration was efficient to transfer a phosphate group from acetyl phosphate to dGR. After 12 h of incubation, a maximal dGMP yield of up to 76% was obtained based on the addition of 5 mM guanosine and 5 mM thymidine.
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KEYWORD
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2′-deoxyguanylic acid, N-deoxyribosyltransferase, purine nucleoside phosphorylase, deoxyguanosine kinase, acetate kinase
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