One-pot multi-enzymatic production of purine derivatives with application in pharmaceutical and food industry
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Abstract
Biocatalysis reproduce nature’s synthetic strategies in order to synthesize different organic compounds. Natural metabolic pathways usually involve complex networks to support cellular growth and survival. In this regard, multi-enzymatic systems are valuable tools for the production of a wide variety of organic compounds. Methods: The production of different purine nucleosides and nucleoside-50 -monophosphates has been performed for first time, catalyzed by the sequential action of 2 0 -deoxyribosyltransferase from Lactobacillus delbrueckii (LdNDT) and hypoxanthine-guanine-xanthine phosphoribosyltransferase from Thermus themophilus HB8 (TtHGXPRT). Results: The biochemical characterization of LdNDT reveals that the enzyme is active and stable in a broad range of pH, temperature, and ionic strength. Substrate specificity studies showed a high promiscuity in the recognition of purine analogues. Finally, the enzymatic production of different purine derivatives was performed to evaluate the efficiency of multi-enzymatic system LdNDT/TtHGXPRT. Conclusions: The production of different therapeutic purine nucleosides was efficiently catalyzed by LdNDT/TtHGXPRT. In addition, the resulting by-products were converted to IMP and GMP. Taking all of these features, this bioprocess entails an efficient, sustainable, and economical alternative to chemical synthetic methods.
Referencias:
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1. Parker, W.B. Enzymology of purine and pyrimidine antimetabolites used in the treatment of cancer. Chem. Rev. 2009, 109, 2880–2893. [CrossRef] [PubMed] 2. De Clercq, E. Recent highlights in the development of new antiviral drugs. Curr. Opin. Microbiol. 2005, 8, 552–560. [CrossRef] [PubMed] 3. Behrens, M.; Meyerhof, W.; Hellfritsch, C.; Hofmann, T. Sweet and umami taste: Natural products, their chemosensory targets, and beyond. Angew. Chem. Int. Ed. 2011, 50, 2220–2242. [CrossRef] [PubMed] 4. Mikhailopulo, I.A. Biotechnology of nucleic acid constituents—State of the art and perspectives. Curr. Org. Chem. 2007, 11, 317–335. [CrossRef] 5. Fresco-Taboada, A.; de la Mata, I.; Arroyo, M.; Fernández-Lucas, J. New insights on nucleoside 2 0 -deoxyribosyltransferases: A versatile biocatalyst for one-pot one-step synthesis of nucleoside analogs. Appl. Microbiol. Biotechnol. 2013, 97, 3773–3785. [CrossRef] [PubMed] 6. Lapponi, M.J.; Rivero, C.W.; Zinni, M.A.; Britos, C.N.; Trelles, J.A. New developments in nucleoside analogues biosynthesis: A review. J. Mol. Catal. B Enzym. 2016, 133, 218–233. [CrossRef] 7. Del Arco, J.; Fernández-Lucas, J. Purine and Pyrimidine Phosphoribosytransferases: A versatile tool for enzymatic synthesis of nucleoside-50 -monophosphates. Curr. Pharm. Des. 2017, 23. [CrossRef] 8. Lewkowicz, E.S.; Iribarren, A.M. Nucleoside phosphorylases. Curr. Org. Chem. 2006, 10, 1197–1215. [CrossRef] 9. Del Arco, J.; Acosta, J.; Pereira, H.M.; Perona, A.; Lokanath, N.K.; Kunishima, N.; Fernández-Lucas, J. Enzymatic production of non-natural nucleoside-50 -monophosphates by a novel thermostable uracil phosphoribosyltransferase. ChemCatChem 2017. [CrossRef] 10. Del Arco, J.; Martinez, M.; Donday, M.; Clemente-Suarez, V.J.; Fernández-Lucas, J. Cloning, expression and biochemical characterization of xanthine and adenine phosphoribosyltransferases from Thermus thermophilus HB8. Biocatal. Biotransfor. 2017, 1–8. [CrossRef] 11. Del Arco, J.; Cejudo-Sanches, J.; Esteban, I.; Clemente-Suarez, V.J.; Hormigo-Cisneros, D.; Perona, A.; Fernández-Lucas, J. Enzymatic production of dietary nucleotides from low-soluble purine bases by an efficient, thermostable and alkali-tolerant biocatalyst. Food Chem. 2017, 237, 605–611. [CrossRef] [PubMed] 12. Serra, I.; Conti, S.; Piškur, J.; Clausen, A.R.; Munch-Petersen, B.; Terreni, M.; Ubiali, D. Immobilized Drosophila melanogaster deoxyribonucleoside kinase (DmdNK) as a high performing biocatalyst for the synthesis of purine arabinonucleotides. Adv. Synth. Catal. 2014, 356, 563–570. [CrossRef] 13. Fernández-Lucas, J. Multienzymatic synthesis of nucleic acid derivatives: A general perspective. Appl. Microbiol. Biotechnol. 2015, 99, 4615–4627. [CrossRef] [PubMed] 14. Zou, Z.; Ding, Q.; Ou, L.; Yan, B. Efficient production of deoxynucleoside-50 -monophosphates using deoxynucleoside kinase coupled with a GTP-regeneration system. Appl. Microbiol. Biotechnol. 2013, 97, 9389–9395. [CrossRef] [PubMed] 15. Mori, H.; Iida, A.; Fujio, T.; Teshiba, S. A novel process of inosine 50 -monophosphate production using overexpressed guanosine/inosine kinase. Appl. Microbiol. Biotechnol. 1997, 48, 693–698. [CrossRef] [PubMed] 16. Zhou, X.; Szeker, K.; Janocha, B.; Böhme, T.; Albrecht, D.; Mikhailopulo, I.A.; Neubauer, P. Recombinant purine nucleoside phosphorylases from thermophiles: Preparation, properties and activity towards purine and pyrimidine nucleosides. FEBS J. 2013, 280, 1475–1490. [CrossRef] [PubMed] 17. Iglesias, L.E.; Lewkowicz, E.S.; Medici, R.; Bianchi, P.; Iribarren, A.M. Biocatalytic approaches applied to the synthesis of nucleoside prodrugs. Biotechnol. Adv. 2015, 33, 412–434. [CrossRef] [PubMed] 18. Fernández-Lucas, J.; Acebal, C.; Sinisterra, J.V.; Arroyo, M.; de la Mata, I. Lactobacillus reuteri 2’-deoxyribosyltransferase, a novel biocatalyst for tailoring of nucleosides. Appl. Environ. Microbiol. 2010, 76, 1462–1470. [CrossRef] [PubMed] 19. Crespo, N.; Sánchez-Murcia, P.A.; Gago, F.; Cejudo-Sanches, J.; Galmes, M.A.; Fernández-Lucas, J.; Mancheño, J.M. 20 -Deoxyribosyltransferase from Leishmania mexicana, an efficient biocatalyst for one-pot, one-step synthesis of nucleosides from poorly soluble purine bases. Appl. Microbiol. Biotechnol. 2017, 101, 7187–7200. [CrossRef] [PubMed] 20. Fresco-Taboada, A.; Serra, I.; Arroyo, M.; Fernández-Lucas, J.; de la Mata, I.; Terreni, M. Development of an immobilized biocatalyst based on Bacillus psychrosaccharolyticus NDT for the preparative synthesis of trifluridine and decytabine. Catal. Today 2016, 259, 197–204. [CrossRef] 21. De Clercq, E. Highlights in the discovery of antiviral drugs: A personal retrospective. J. Med. Chem. 2010, 53, 1438–1450. [CrossRef] [PubMed] 22. Mateo, C.; Palomo, J.M.; Fernandez-Lorente, G.; Guisan, J.M.; Fernandez-Lafuente, R. Improvement of enzyme activity, stability and selectivity via immobilization techniques. Enzym. Microb. Technol. 2007, 40, 1451–1463. [CrossRef] 23. Laemmli, U.K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970, 227, 680–685. [CrossRef] [PubMed] 24. Gill, S.C.; Von Hippel, P.H. Calculation of protein extinction coefficients from amino acid sequence data. Anal. Biochem. 1989, 182, 319–326. [CrossRef]
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