title: Efficient production of deoxynucleoside-5'-monophosphates using deoxynucleoside kinase coupled with a GTP-regeneration system.
abstract: Deoxynucleoside-5'-monophosphates (5'-dNMPs) are the basic components of DNA and are widely used in medicine and as chemical and biochemical reagents. A large amount of effort has been expended to obtain 5'-dNMPs of high quality and at a low cost. However, these procedures are inefficient and inconvenient. In this study, deoxyadenosine-5'-monophosphate (5'-dAMP), 2,6-diaminopurine deoxynucleoside-5'-monophosphate (5'-dDAMP), and deoxycytidine-5'-monophosphate (5'-dCMP) were biosynthesized using recombinant N-deoxyribosyltransferase II (NDT-II), deoxycytidine kinase, and acetate kinase in a one-pot reaction system. The ndt-II gene from Lactobacillus delbrueckii, dck from Bacillus subtilus, and ack from Escherichia coli K12 were overexpressed in E. coli BL21 (DE3). Thymidine was used as the deoxyribose donor; GTP was used as the phosphate donor, and acetyl phosphate was used to regenerate GTP. Under optimized conditions, each 10 mM adenine, 10 mM 2,6-diaminopurine, or 10 mM cytosine were converted into 9.01 mM 5'-dAMP, 8.68 mM 5'-dDAMP, or 6.23 mM 5'-dCMP, respectively. The high yield indicated that this process of biosynthesis of 5'-dAMP, 5'-dDAMP, or 5'-dCMP was efficient and economical, and this one-pot system may also potentially be used for the preparation of other types of 5'-dNMPs.
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China.
authors: Zhi
authors: Z
authors: Zou
authors: Qingbao
authors: Q
authors: Ding
authors: Ling
authors: L
authors: Ou
authors: Bingkun
authors: B
authors: Yan
mesh_terms: D001407:Bacillus / Q000201:enzymology / Q000235:genetics; D003067:Coenzymes / Q000378:metabolism*; D003838:Deoxyadenine Nucleotides / Q000378:metabolism*; D003843:Deoxycytidine Monophosphate / Q000378:metabolism*; D048168:Escherichia coli K12 / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression; D006160:Guanosine Triphosphate / Q000378:metabolism*; D048668:Lactobacillus delbrueckii / Q000201:enzymology / Q000235:genetics; D060847:Metabolic Engineering; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D003067:Coenzymes; D003838:Deoxyadenine Nucleotides; D011994:Recombinant Proteins; D003843:Deoxycytidine Monophosphate; C012873:2'-deoxy-5'-adenosine monophosphate; D006160:Guanosine Triphosphate; D017853:Phosphotransferases (Alcohol Group Acceptor); C020018:deoxyribonucleoside kinases