Abstract
2′-Deoxyribonucleosides are important as building blocks for the synthesis of antisense drugs, antiviral nucleosides, and 2′-deoxyribonucleotides for polymerase chain reaction. The microbial production of 2′-deoxyribonucleosides from simple materials, glucose, acetaldehyde, and a nucleobase, through the reverse reactions of 2′-deoxyribonucleoside degradation and the glycolytic pathway, was investigated. The glycolytic pathway of baker’s yeast yielded fructose 1,6-diphosphate from glucose using the energy of adenosine 5′-triphosphate generated from adenosine 5′-monophosphate through alcoholic fermentation with the yeast. Fructose 1,6-diphosphate was further transformed to 2-deoxyribose 5-phosphate in the presence of acetaldehyde by deoxyriboaldolase-expressing Escherichia coli cells via d-glyceraldehyde 3-phosphate. E. coli transformants expressing phosphopentomutase and nucleoside phosphorylase produced 2′-deoxyribonucleosides from 2-deoxyribose 5-phosphate and a nucleobase via 2-deoxyribose 1-phosphate through the reverse reactions of 2′-deoxyribonucleoside degradation. Coupling of the glycolytic pathway and deoxyriboaldolase-catalyzing reaction efficiently supplied 2-deoxyribose 5-phosphate, which is a key intermediate for 2′-deoxyribonucleoside synthesis. 2′-Deoxyinosine (9.9 mM) was produced from glucose, acetaldehyde, and adenine through three-step reactions via fructose 1,6-diphosphate and then 2-deoxyribose 5-phosphate, the molar yield as to glucose being 17.8%.
Similar content being viewed by others
References
Aoyama H (1987) Stereoselective synthesis of anomers of 5-substituted 2′-deoxyuridines. Bull Chem Soc Jpn 60:2073–2077
Brunella A, Ghisalba O (2000) Recombinant Lactobacillus leichmannii ribonucleoside triphosphate reductase as biocatalyst in the preparative synthesis of 2′-deoxyribonucleoside-5′-triphosphates. J Mol Catal B Enzym 10:215–222
Brunella A, Abrantes M, Ghisalba O (2000) Preparative 2′-reduction of ATP catalyzed by ribonucleotide reductase purified by liquid–liquid extraction. Biosci Biotechnol Biochem 64:1836–1841
Fischer M, Short SA (1982) The cloning of the Escherichia coli K-12 deoxyribonucleoside operon. Gene 17:291–298
Fruya A, Kato F, Nakayama K (1975) Accumulation of 6-azauridine by mutants of Brevibacterium ammoniagenes. Agric Biol Chem 39:767–771
Fujio T, Maruyama A (1997) Enzymatic production of pyrimidine nucleotides using Corynebacterium ammoniagenes cells and recombinant Escherichia coli cells: enzymatic production of CDP-choline from orotic acid and choline chloride (part I). Biosci Biotechnol Biochem 61:956–959
Harden A, Young W (1905) The alcoholic fermentation of yeast-juice. Proc R Soc Lond B Biol Sci 77:405–420
Hoffer M (1960) α-Thymidine. Chem Ber 93:2777–2781
Horinouchi N, Ogawa J, Sakai T, Kawano T, Matsumoto S, Sasaki M, Mikami Y, Shimizu S (2003) Construction of deoxyriboaldolase-expressing Escherichia coli and its application to 2-deoxyribose 5-phosphate synthesis from glucose and acetaldehyde for 2’-deoxyribonucleoside production. Appl Environ Microbiol 69:3791–3797
Ishii K, Shiio I (1972) Improved inosine production and derepression of purine nucleotide biosynthetic enzymes in 8-azaguanine resistant mutants of Bacillus subtilis. Agric Biol Chem 36:1511–1522
Kawakami H, Matsushita H, Naoi Y, Itoh K, Yoshikoshi H (1989) The synthesis of 2′-deoxyadenosine via stereospecific coupling reaction. Chem Lett 235–238
Ogata K (1975) The microbial production of nucleic acid-related compounds. Adv Appl Microbiol 19:209–247
Ogawa J, Saito K, Sakai T, Horinouchi N, Kawano T, Matsumoto S, Sasaki M, Mikami Y, Shimizu S (2003) Microbial production of 2-deoxyribose 5-phosphate from acetaldehyde and triosephosphate for the synthesis of 2′-deoxyribonucleosides. Biosci Biotechnol Biochem 67:933–936
Ouwerkerk N, van Boom JH, Lugtenburg J, Raap J (2000) Chemo-enzymatic synthesis of thymidine 13C-labelled in the 2′-deoxyribose moiety. Eur J Org Chem 861–866
Ouwerkerk N, Steenweg M, de Ruijter M, Brouwer J, van Boom JH, Lugtenburg J, Raap J (2002) One-pot two-step enzymatic coupling of pyrimidine bases to 2-deoxy-D-ribose-5-phosphate. A new strategy in the synthesis of stable isotope labeled deoxynucleosides. J Org Chem 67:1480–1489
Park M, Rizzo CJ (1996) Stereocontrolled de novo synthesis of β-2′-deoxynucleosides. J Org Chem 61:6092–6093
Prazeres DMF, Ferreira GNM, Monterio GA, Cooney CL, Cabral JMS (1999) Large-scale production of pharmaceutical-grade plasmid DNA for gene therapy: problems and bottlenecks. Trends Biotechnol 17:169–174
Satoh T, Ishizaki A, Ueda S (1989) Production of DNA by continuous culture using Pseudomonas aeruginosa KYU-1. J Ferment Bioeng 68:92–95
Schmid A, Dordick JS, Hauer B, Kiener A, Wubbolts M, Witholt B (2001) Industrial biocatalysis today and tomorrow. Nature 409:258–268
Stumpf PK (1947) A colorimetric method for the determination of deoxyribonucleic acid. J Biol Chem 169:367–371
Tomita F, Suzuki T (1972) Extracellular accumulation of DNA by hydrocarbon-utilizing bacteria. Agric Biol Chem 36:133–140
Utagawa T (1999) Enzymatic production of nucleoside antibiotics. J Mol Catal B Enzym 6:215–222
Wakisaka S, Ohshima Y, Ogawa M, Tochikura T, Tachiki T (1998) Characteristics and efficiency of glutamine production by coupling of a bacterial glutamine synthetase reaction with the alcoholic fermentation system of baker’s yeast. Appl Environ Microbiol 64:2953–2957
Yokozeki K, Tsuji T (2000) A novel enzymatic method for the production of purine-2′-deoxyribonucleosides. J Mol Catal B Enzym 10:207–213
Acknowledgements
This work was partially supported by the Industrial Technology Research Grant Program (No. 02A07001a to J.O.) and the Project for Development of a Technological Infrastructure for Industrial Bioprocesses on R&D of New Industrial Science and Technology Frontiers (to S.S.) of the New Energy and Industrial Technology Development Organization, Japan, and Grants-in-Aid for Scientific Research (No. 16688004 to J.O.) and COE for Microbial-Process Development Pioneering Future Production Systems (to S. S.) from the Ministry of Education, Science, Sports, and Culture, Japan.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Horinouchi, N., Ogawa, J., Kawano, T. et al. Biochemical retrosynthesis of 2′-deoxyribonucleosides from glucose, acetaldehyde, and a nucleobase. Appl Microbiol Biotechnol 71, 615–621 (2006). https://doi.org/10.1007/s00253-005-0205-5
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-005-0205-5